Improved in vitro models for preclinical drug and formulation screening focusing on 2D and 3D skin and cornea constructs.

PubMed

Beißner, Nicole; Bolea Albero, Antonio; Füller, Jendrik; Kellner, Thomas; Lauterboeck, Lothar; Liang, Jinghu; Böl, Markus; Glasmacher, Birgit; Müller-Goymann, Christel C; Reichl, Stephan

2018-05-01

The present overview deals with current approaches for the improvement of in vitro models for preclinical drug and formulation screening which were elaborated in a joint project at the Center of Pharmaceutical Engineering of the TU Braunschweig. Within this project a special focus was laid on the enhancement of skin and cornea models. For this reason, first, a computation-based approach for in silico modeling of dermal cell proliferation and differentiation was developed. The simulation should for example enhance the understanding of the performed 2D in vitro tests on the antiproliferative effect of hyperforin. A second approach aimed at establishing in vivo-like dynamic conditions in in vitro drug absorption studies in contrast to the commonly used static conditions. The reported Dynamic Micro Tissue Engineering System (DynaMiTES) combines the advantages of in vitro cell culture models and microfluidic systems for the emulation of dynamic drug absorption at different physiological barriers and, later, for the investigation of dynamic culture conditions. Finally, cryopreserved shipping was investigated for a human hemicornea construct. As the implementation of a tissue-engineering laboratory is time-consuming and cost-intensive, commercial availability of advanced 3D human tissue is preferred from a variety of companies. However, for shipping purposes cryopreservation is a challenge to maintain the same quality and performance of the tissue in the laboratory of both, the provider and the customer. Copyright © 2017 Elsevier B.V. All rights reserved.

Silk fibroin in tissue engineering.

PubMed

Kasoju, Naresh; Bora, Utpal

2012-07-01

Tissue engineering (TE) is a multidisciplinary field that aims at the in vitro engineering of tissues and organs by integrating science and technology of cells, materials and biochemical factors. Mimicking the natural extracellular matrix is one of the critical and challenging technological barriers, for which scaffold engineering has become a prime focus of research within the field of TE. Amongst the variety of materials tested, silk fibroin (SF) is increasingly being recognized as a promising material for scaffold fabrication. Ease of processing, excellent biocompatibility, remarkable mechanical properties and tailorable degradability of SF has been explored for fabrication of various articles such as films, porous matrices, hydrogels, nonwoven mats, etc., and has been investigated for use in various TE applications, including bone, tendon, ligament, cartilage, skin, liver, trachea, nerve, cornea, eardrum, dental, bladder, etc. The current review extensively covers the progress made in the SF-based in vitro engineering and regeneration of various human tissues and identifies opportunities for further development of this field. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The Application of Sheet Technology in Cartilage Tissue Engineering.

PubMed

Ge, Yang; Gong, Yi Yi; Xu, Zhiwei; Lu, Yanan; Fu, Wei

2016-04-01

Cartilage tissue engineering started to act as a promising, even essential alternative method in the process of cartilage repair and regeneration, considering adult avascular structure has very limited self-renewal capacity of cartilage tissue in adults and a bottle-neck existed in conventional surgical treatment methods. Recent progressions in tissue engineering realized the development of more feasible strategies to treat cartilage disorders. Of these strategies, cell sheet technology has shown great clinical potentials in the regenerative areas such as cornea and esophagus and is increasingly considered as a potential way to reconstruct cartilage tissues for its non-use of scaffolds and no destruction of matrix secreted by cultured cells. Acellular matrix sheet technologies utilized in cartilage tissue engineering, with a sandwich model, can ingeniously overcome the drawbacks that occurred in a conventional acellular block, where cells are often blocked from migrating because of the non-nanoporous structure. Electrospun-based sheets with nanostructures that mimic the natural cartilage matrix offer a level of control as well as manipulation and make them appealing and widely used in cartilage tissue engineering. In this review, we focus on the utilization of these novel and promising sheet technologies to construct cartilage tissues with practical and beneficial functions.

[The 2009 performance report of the German cornea banks].

PubMed

Schrage, N; Reinhard, T; Seitz, B; Hermel, M; Böhringer, D; Reinshagen, H

2011-03-01

In Germany, human tissue for corneal and amniotic transplantation is supplied by 27 cornea banks. The Section for Tissue Transplantation and Biotechnology of the German Ophthalmological Society records the cornea banks' activities by means of an annual questionnaire. In 2009, a total of 4,818 corneal grafts were processed by 21 responding cornea banks, and 57% were deemed suitable for transplantation. This ratio is slightly higher than the European average. In addition, German cornea banks released 1,257 amniotic grafts in 2009. German cornea banks are currently facing new regulatory issues due to updated legislation regarding tissue transplantation. Recent updates in European law have limited the cutoff time for postmortem blood sampling to 24 h, and this regulation may lead to a significant reduction in potential donors.

Cornea Transplant

MedlinePlus

... transplants may be used that remove only certain layers of cornea tissue, or only tissue affected by ... procedure removes diseased tissue from the back corneal layers, including the endothelium, along with the Descemet membrane, ...

Lactoferrin Expression in Human and Murine Ocular Tissue.

PubMed

Rageh, Abrar A; Ferrington, Deborah A; Roehrich, Heidi; Yuan, Ching; Terluk, Marcia R; Nelson, Elizabeth F; Montezuma, Sandra R

2016-07-01

Lactoferrin (LF) is a multifunctional protein known to provide innate defense due to its antimicrobial and anti-inflammatory properties. In the eye, LF has been identified in the tears and vitreous humor. Its presence in other ocular tissues has not been determined. Our aim is to assess the presence of LF in the cornea, iris, retina and retinal pigment epithelium (RPE) of humans and mice. To test for the endogenous production of LF, reverse transcription polymerase chain reaction was performed in cultured human cells from the cornea and RPE and in murine tissues. To confirm LF localization in specific ocular tissue, immunohistochemistry was performed on flat mounts of cornea, retina and RPE in human donor eyes. The presence of LF was assessed by western blotting in human and mouse ocular tissue and human culture cells (cornea and RPE). To verify antibody specificity, purified human LF and transferrin (TF) were used on 1D and 2D western blots. LF gene expression was confirmed in the cornea and RPE cell cultures from humans, suggesting that LF is an endogenously produced protein. PCR results from mouse ocular tissue showed LF expression in cornea, iris, RPE, but not in retina. These results were also consistent with immunohistochemical localization of LF in human donor tissue. Antibody reaction for human LF was specific and western blotting showed its presence in the cornea, iris and RPE tissues. A faint reaction for the retina was observed but was likely due to contamination from other ocular tissues. Multiple commercially available antibodies for murine LF cross-reacted with TF, so no reliable results were obtained for murine western blot. LF is expressed in multiple eye tissues of humans and mice. This widespread expression and multifunctional activity of LF suggests that it may play an important role in protecting eye tissues from inflammation-associated diseases.

Topographical Control of Ocular Cell Types for Tissue Engineering

PubMed Central

McHugh, Kevin J.; Saint-Geniez, Magali; Tao, Sarah L.

2014-01-01

Visual impairment affects over 285 million people worldwide and has a major impact on an individual’s quality of life. Tissue engineering has the potential to increase quality of life for many of these patients by preventing vision loss or restoring vision using cell-based therapies. However, these strategies will require an understanding of the microenvironmental factors that influence cell behavior. The eye is a well-organized organ whose structural complexity is essential for proper function. Interactions between ocular cells and their highly ordered extracellular matrix are necessary for maintaining key tissue properties including corneal transparency and retinal lamination. Therefore, it is not surprising that culturing these cells in vitro on traditional flat substrates result in irregular morphology. Instead, topographically patterned biomaterials better mimic native extracellular matrix and have been shown to elicit in vivo-like morphology and gene expression which is essential for tissue engineering. Herein we review multiple methods for producing well-controlled topography and discuss optimal biomaterial scaffold design for cells of the cornea, retina, and lens. PMID:23744715

Wound-Healing Studies in Cornea and Skin: Parallels, Differences and Opportunities

PubMed Central

Bukowiecki, Anne; Hos, Deniz; Cursiefen, Claus; Eming, Sabine A.

2017-01-01

The cornea and the skin are both organs that provide the outer barrier of the body. Both tissues have developed intrinsic mechanisms that protect the organism from a wide range of external threats, but at the same time also enable rapid restoration of tissue integrity and organ-specific function. The easy accessibility makes the skin an attractive model system to study tissue damage and repair. Findings from skin research have contributed to unravelling novel fundamental principles in regenerative biology and the repair of other epithelial-mesenchymal tissues, such as the cornea. Following barrier disruption, the influx of inflammatory cells, myofibroblast differentiation, extracellular matrix synthesis and scar formation present parallel repair mechanisms in cornea and skin wound healing. Yet, capillary sprouting, while pivotal in proper skin wound healing, is a process that is rather associated with pathological repair of the cornea. Understanding the parallels and differences of the cellular and molecular networks that coordinate the wound healing response in skin and cornea are likely of mutual importance for both organs with regard to the development of regenerative therapies and understanding of the disease pathologies that affect epithelial-mesenchymal interactions. Here, we review the principal events in corneal wound healing and the mechanisms to restore corneal transparency and barrier function. We also refer to skin repair mechanisms and their potential implications for regenerative processes in the cornea. PMID:28604651

Wound-Healing Studies in Cornea and Skin: Parallels, Differences and Opportunities.

PubMed

Bukowiecki, Anne; Hos, Deniz; Cursiefen, Claus; Eming, Sabine A

2017-06-12

The cornea and the skin are both organs that provide the outer barrier of the body. Both tissues have developed intrinsic mechanisms that protect the organism from a wide range of external threats, but at the same time also enable rapid restoration of tissue integrity and organ-specific function. The easy accessibility makes the skin an attractive model system to study tissue damage and repair. Findings from skin research have contributed to unravelling novel fundamental principles in regenerative biology and the repair of other epithelial-mesenchymal tissues, such as the cornea. Following barrier disruption, the influx of inflammatory cells, myofibroblast differentiation, extracellular matrix synthesis and scar formation present parallel repair mechanisms in cornea and skin wound healing. Yet, capillary sprouting, while pivotal in proper skin wound healing, is a process that is rather associated with pathological repair of the cornea. Understanding the parallels and differences of the cellular and molecular networks that coordinate the wound healing response in skin and cornea are likely of mutual importance for both organs with regard to the development of regenerative therapies and understanding of the disease pathologies that affect epithelial-mesenchymal interactions. Here, we review the principal events in corneal wound healing and the mechanisms to restore corneal transparency and barrier function. We also refer to skin repair mechanisms and their potential implications for regenerative processes in the cornea.

Nerve Repulsion by the Lens and Cornea during Cornea Innervation is Dependent on Robo-Slit Signaling and Diminishes with Neuron Age

PubMed Central

Schwend, Tyler; Lwigale, Peter Y.; Conrad, Gary W.

2012-01-01

The cornea, the most densely innervated tissue on the surface of the body, becomes innervated in a series of highly coordinated developmental events. During cornea development, chick trigeminal nerve growth cones reach the cornea margin at embryonic day (E)5, where they are initially repelled for days from E5-8, instead encircling the corneal periphery in a nerve ring prior to entering on E9. The molecular events coordinating growth cone guidance during cornea development are poorly understood. Here we evaluated a potential role for the Robo-Slit nerve guidance family. We found that Slit 1, 2 and 3 expression in the cornea and lens persisted during all stages of cornea innervation examined. Robo1 expression was developmentally regulated in trigeminal cell bodies, expressed robustly during nerve ring formation (E5-8), then later declining concurrent with projection of growth cones into the cornea. In this study we provide in vivo and in vitro evidence that Robo-Slit signaling guides trigeminal nerves during cornea innervation. Transient, localized inhibition of Robo-Slit signaling, by means of beads loaded with inhibitory Robo-Fc protein implanted into the developing eyefield in vivo, led to disorganized nerve ring formation and premature cornea innervation. Additionally, when trigeminal explants (source of neurons) were oriented adjacent to lens vesicles or corneas (source of repellant molecules) in organotypic tissue culture both lens and cornea tissues strongly repelled E7 trigeminal neurites, except in the presence of inhibitory Robo-Fc protein. In contrast, E10 trigeminal neurites were not as strongly repelled by cornea, and presence of Robo-Slit inhibitory protein had no effect. In full, these findings suggest that nerve repulsion from the lens and cornea during nerve ring formation is mediated by Robo-Slit signaling. Later, a shift in nerve guidance behavior occurs, in part due to molecular changes in trigeminal neurons, including Robo1 downregulation, thus allowing nerves to find the Slit-expressing cornea permissive for growth cones. PMID:22236962

Diclofenac sodium delivery to the eye: in vitro evaluation of novel solid lipid nanoparticle formulation using human cornea construct.

PubMed

Attama, Anthony A; Reichl, Stephan; Müller-Goymann, Christel C

2008-05-01

Solid lipid nanoparticles (SLNs) were prepared with a combination of homolipid from goat (goat fat) and phospholipid, and evaluated for diclofenac sodium (DNa) delivery to the eye using bio-engineered human cornea, produced from immortalized human corneal endothelial cells (HENC), stromal fibroblasts and epithelial cells CEPI 17 CL 4. Encapsulation efficiency was high and sustained release of DNa and high permeation through the bio-engineered cornea were achieved. Results obtained in this work showed that permeation of DNa through the cornea construct was improved by formulation as SLN modified with phospholipid.

The Resin-Embedded Cornea Prepared Via Rapid Processing Protocol : A Good Histomorphometric Target for Clinical Investigation in Ophthalmology and Optometry

PubMed Central

Cheah, Pike See; Mohidin, Norhani; Mohd Ali, Bariah; Maung, Myint; Latif, Azian Abdul

2008-01-01

This study illustrates and quantifies the changes on corneal tissue between the paraffin-embedded and resin-embedded blocks and thus, selects a better target in investigational ophthalmology and optometry via light microscopy. Corneas of two cynomolgus monkeys (Macaca fascicularis) were used in this study. The formalin-fixed cornea was prepared in paraffin block via the conventional tissue processing protocol (4-day protocol) and stained with haematoxylin and eosin. The glutaraldehyde-fixed cornea was prepared in resin block via the rapid and modified tissue processing procedure (1.2-day protocol) and stained with toluidine blue. The paraffin-embedded sample exhibits various undesired tissue damage and artifact such as thinner epithelium (due to the substantial volumic extraction from the tissue), thicker stroma layer (due to the separation of lamellae and the presence of voids) and the distorted endothelium. In contrast, the resin-embedded corneal tissue has demonstrated satisfactory corneal ultrastructural preservation. The rapid and modified tissue processing method for preparing the resin-embedded is particularly beneficial to accelerate the microscopic evaluation in ophthalmology and optometry. PMID:22570589

Improving the moisturizing properties of collagen film by surface grafting of chondroitin sulfate for corneal tissue engineering.

PubMed

Liu, Yang; Lv, Huilin; Ren, Li; Xue, Guanhua; Wang, Yingjun

2016-01-01

Cornea disease is the second cause of blindness and keratoplasty is the most commonly performed option for visual rehabilitation of patients with corneal blindness. However, the clinical treatment has been drastically limited due to a severe shortage of high-quality donor corneas. Although collagen film with outstanding biocompatibility has promising application in corneal tissue engineering, the moisturizing properties of collagen-based materials must be further improved to satisfy the requirements of clinical applications. This paper describes a novel collagen-based film with high moisture capacity reinforced by surface grafting of chondroitin sulfate. The collagen-chondroitin sulfate (abbreviated as Col-CS) film was analyzed by Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy and its hydrophilic property, moisture retention, optical property, and mechanical performance had been tested. The moisture-retaining capacity is found to be improved with the introduction of chondroitin sulfate, and the Col-CS membrane performs better mechanical properties than the collagen film. Moreover, the modified film proves excellent biocompatibility for the proliferation of human corneal epithelial cells in vitro. This Col-CS film with good moisturizing properties can reduce the risk of xerophthalmia and is expected to increase the implant success rate in clinic patients with corneal defects.

Expansion and cryopreservation of porcine and human corneal endothelial cells.

PubMed

Marquez-Curtis, Leah A; McGann, Locksley E; Elliott, Janet A W

2017-08-01

Impairment of the corneal endothelium causes blindness that afflicts millions worldwide and constitutes the most often cited indication for corneal transplants. The scarcity of donor corneas has prompted the alternative use of tissue-engineered grafts which requires the ex vivo expansion and cryopreservation of corneal endothelial cells. The aims of this study are to culture and identify the conditions that will yield viable and functional corneal endothelial cells after cryopreservation. Previously, using human umbilical vein endothelial cells (HUVECs), we employed a systematic approach to optimize the post-thaw recovery of cells with high membrane integrity and functionality. Here, we investigated whether improved protocols for HUVECs translate to the cryopreservation of corneal endothelial cells, despite the differences in function and embryonic origin of these cell types. First, we isolated endothelial cells from pig corneas and then applied an interrupted slow cooling protocol in the presence of dimethyl sulfoxide (Me 2 SO), with or without hydroxyethyl starch (HES). Next, we isolated and expanded endothelial cells from human corneas and applied the best protocol verified using porcine cells. We found that slow cooling at 1 °C/min in the presence of 5% Me 2 SO and 6% HES, followed by rapid thawing after liquid nitrogen storage, yields membrane-intact cells that could form monolayers expressing the tight junction marker ZO-1 and cytoskeleton F-actin, and could form tubes in reconstituted basement membrane matrix. Thus, we show that a cryopreservation protocol optimized for HUVECs can be applied successfully to corneal endothelial cells, and this could provide a means to address the need for off-the-shelf cryopreserved cells for corneal tissue engineering and regenerative medicine. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

A simplified technique for in situ excision of cornea and evisceration of retinal tissue from human ocular globe.

PubMed

Parekh, Mohit; Ferrari, Stefano; Di Iorio, Enzo; Barbaro, Vanessa; Camposampiero, Davide; Karali, Marianthi; Ponzin, Diego; Salvalaio, Gianni

2012-06-12

Enucleation is the process of retrieving the ocular globe from a cadaveric donor leaving the rest of the globe undisturbed. Excision refers to the retrieval of ocular tissues, especially cornea, by cutting it separate from the ocular globe. Evisceration is the process of removing the internal organs referred here as retina. The ocular globe consists of the cornea, the sclera, the vitreous body, the lens, the iris, the retina, the choroid, muscles etc (Suppl. Figure 1). When a patient is suffering from corneal damage, the cornea needs to be removed and a healthy one must be transplanted by keratoplastic surgeries. Genetic disorders or defects in retinal function can compromise vision. Human ocular globes can be used for various surgical procedures such as eye banking, transplantation of human cornea or sclera and research on ocular tissues. However, there is little information available on human corneal and retinal excision, probably due to the limited accessibility to human tissues. Most of the studies describing similar procedures are performed on animal models. Research scientists rely on the availability of properly dissected and well-conserved ocular tissues in order to extend the knowledge on human eye development, homeostasis and function. As we receive high amount of ocular globes out of which approximately 40% (Table 1) of them are used for research purposes, we are able to perform huge amount of experiments on these tissues, defining techniques to excise and preserve them regularly. The cornea is an avascular tissue which enables the transmission of light onto the retina and for this purpose should always maintain a good degree of transparency. Within the cornea, the limbus region, which is a reservoir of the stem cells, helps the reconstruction of epithelial cells and restricts the overgrowth of the conjunctiva maintaining corneal transparency and clarity. The size and thickness of the cornea are critical for clear vision, as changes in either of them could lead to distracted, unclear vision. The cornea comprises of 5 layers; a) epithelium, b) Bowman's layer, c) stroma, d) Descemet's membrane and e) endothelium. All layers should function properly to ensure clear vision(4,5,6). The choroid is the intermediate tunic between the sclera and retina, bounded on the interior by the Bruch's membrane and is responsible for blood flow in the eye. The choroid also helps to regulate the temperature and supplies nourishment to the outer layers of the retina(5,6). The retina is a layer of nervous tissue that covers the back of the ocular globe (Suppl. Figure 1) and consists of two parts: a photoreceptive part and a non-receptive part. The retina helps to receive the light from the cornea and lens and converts it into the chemical energy eventually transmitted to the brain with help of the optic nerve(5,6). The aim of this paper is to provide a protocol for the dissection of corneal and retinal tissues from human ocular globes. Avoiding cross-contamination with adjacent tissues and preserving RNA integrity is of fundamental importance as such tissues are indispensable for research purposes aimed at (i) characterizing the transcriptome of the ocular tissues, (ii) isolating stem cells for regenerative medicine projects, and (iii) evaluating histological differences between tissues from normal/affected subjects. In this paper we describe the technique we currently use to remove the cornea, the choroid and retinal tissues from an ocular globe. Here we provide a detailed protocol for the dissection of the human ocular globe and the excision of corneal and retinal tissues. The accompanying video will help researchers to learn an appropriate technique for the retrieval of precious human tissues which are difficult to find regularly.

Stromal fibroblasts are associated with collagen IV in scar tissues of alkali-burned and lacerated corneas.

PubMed

Ishizaki, M; Shimoda, M; Wakamatsu, K; Ogro, T; Yamanaka, N; Kao, C W; Kao, W W

1997-04-01

Corneal wound healing frequently leads to the formation of opaque scar tissue. We examined whether stromal fibroblastic cells of injured corneas express collagen IV and contributes to the formation of a basal lamina-like structure. Rabbits were anesthetized, and central corneal alkali burn (8 mm in diameter; 1 M NaOH, 1 min) or laceration (8 mm long) were produced. The injured corneas, which had healed for 1, 7, 21 and 45 days, were subjected to histological and immunohistochemical studies with goat anti-collagen IV antibodies, using light and electron microscopy, and in situ hybridization with an antisense digoxigenin-labeled riboprobe of collagen alpha 1(IV) mRNA. For comparison, twenty-day-old fetal corneas were subjected to immunohistochemical study and transmission electron microscopy (TEM). TEM examinations revealed that the stromal collagenous matrix was organized in orthogonal lamellae during corneal development, whereas that of alkali-burned cornea, which had healed for 3 weeks, was disorganized. The stroma of twenty-day-old fetal cornea was not labeled by the anti-collagen IV antibodies. In contrast, one week after injury, specific collagen IV immunostaining was detected in the injured stroma. As the healing proceeded (21-45 days), the antibodies reacted with fibroblastic cells and the extracellular matrix of scar tissues located in the anterior portion of alkali-burned corneas, as well as the posterior portion of lacerated corneas. The middle portion of the stromal tissues was weakly labeled by the anti-collagen IV antibodies with the exception of the blood vessel wall. Immuno-electron microscopic study showed that collagen IV and fibronectin were closely associated with the fibroblastic cells. In situ hybridization demonstrated that epithelial and endothelial cells and fibroblastic cells in the wounded corneal stroma and retro-corneal membrane expressed alpha 1(IV) mRNA, whereas in normal corneas the expression of alpha 1(IV) mRNA was limited to epithelial and endothelial cells. The enhanced expression of collagen IV by the fibroblastic cells in the stroma of injured corneas is consistent with the notion that they may contribute to the formation of basal lamina-like structures in injured corneas.

Covalently immobilized platelet-derived growth factor-BB promotes angiogenesis in biomimetic poly(ethylene glycol) hydrogels

PubMed Central

Saik, Jennifer E.; Gould, Daniel J.; Watkins, Emily M.; Dickinson, Mary E.; West, Jennifer L.

2011-01-01

The field of tissue engineering is severely limited by a lack of microvascularization in tissue engineered constructs. Biomimetic poly(ethylene glycol) hydrogels containing covalently immobilized platelet-derived growth factor BB (PDGF-BB) were developed to promote angiogenesis. Poly(ethylene glycol) hydrogels resist protein absorption and subsequent non-specific cell adhesion, thus providing a “blank slate”, which can be modified through the incorporation of cell adhesive ligands and growth factors. PDGF-BB is a key angiogenic protein able to support neovessel stabilization by inducing functional anastomoses and recruiting pericytes. Due to the widespread effects of PDGF in the body and a half-life of only 30 min in circulating blood, immobilization of PDGF-BB may be necessary. In this work bioactive, covalently immobilized PDGF-BB was shown to induce tubulogenesis on two-dimensional modified surfaces, migration in three-dimensional (3D) degradable hydrogels and angiogenesis in a mouse cornea micro-pocket angiogenesis assay. Covalently immobilized PDGF-BB was also used in combination with covalently immobilized fibroblast growth factor-2, which led to significantly increased endothelial cell migration in 3D degradable hydrogels compared with the presentation of each factor alone. When a co-culture of endothelial cells and mouse pericyte precursor 10T1/2 cells was seeded onto modified surfaces tubule formation was independent of surface modifications with covalently immobilized growth factors. Furthermore, the combination of soluble PDGF-BB and immobilized PDGF-BB induced a more robust vascular response compared with soluble PDGF-BB alone when implanted into an in vivo mouse cornea micropocket angiogenesis assay. Based on these results, we believe bioactive hydrogels can be tailored to improve the formation of functional microvasculature for tissue engineering. PMID:20801242

C3 fixed in vivo to cornea from horses inoculated with Leptospira interrogans.

PubMed

Parma, A E; Cerone, S I; Sansinanea, S A; Ghezzi, M

1992-10-01

C3 was detected bound in vivo to the opaque cornea of horses inoculated with killed Leptospira interrogans. Employing epithelial corneal cells isolated from a monolayer in tissue culture, we proved that C3 is fixed in vitro to the intact cell surface after incubation with a fresh equine anti-Leptospira serum. These findings, in addition to the infiltration of cornea with neutrophils and lymphocytes, may explain the mechanisms of tissue damage in recurrent uveitis of horses with leptospirosis.

Serum response factor: positive and negative regulation of an epithelial gene expression network in the destrin mutant cornea

PubMed Central

Kawakami-Schulz, Sharolyn V.; Verdoni, Angela M.; Sattler, Shannon G.; Jessen, Erik; Kao, Winston W.-Y.; Ikeda, Akihiro

2014-01-01

Increased angiogenesis, inflammation, and proliferation are hallmarks of diseased tissues, and in vivo models of these disease phenotypes can provide insight into disease pathology. Dstncorn1 mice, deficient for the actin depolymerizing factor destrin (DSTN), display an increase of serum response factor (SRF) that results in epithelial hyperproliferation, inflammation, and neovascularization in the cornea. Previous work demonstrated that conditional ablation of Srf from the corneal epithelium of Dstncorn1 mice returns the cornea to a wild-type (WT) like state. This result implicated SRF as a major regulator of genes that contributes to abnormal phenotypes in Dstncorn1 cornea. The purpose of this study is to identify gene networks that are affected by increased expression of Srf in the Dstncorn1 cornea. Microarray analysis led to characterization of gene expression changes that occur when conditional knockout of Srf rescues mutant phenotypes in the cornea of Dstncorn1 mice. Comparison of gene expression values from WT, Dstncorn1 mutant, and Dstncorn1 rescued cornea identified >400 differentially expressed genes that are downstream from SRF. Srf ablation had a significant effect on genes associated with epithelial cell-cell junctions and regulation of actin dynamics. The majority of genes affected by SRF are downregulated in the Dstncorn1 mutant cornea, suggesting that increased SRF negatively affects transcription of SRF gene targets. ChIP-seq analysis on Dstncorn1 mutant and WT tissue revealed that, despite being present in higher abundance, SRF binding is significantly decreased in the Dstncorn1 mutant cornea. This study uses a unique model combining genetic and genomic approaches to identify genes that are regulated by SRF. These findings expand current understanding of the role of SRF in both normal and abnormal tissue homeostasis. PMID:24550211

[Significance of laser confocal tomography in diagnosis and monitoring of keratoconjunctivitis sicca].

PubMed

Safonova, T N; Gladkova, О V; Boev, V I

2016-01-01

Laser confocal tomography of the cornea enables studying ultrathin sections of corneal layers, which provides additional reliable information on tissue changes in keratoconjunctivitis sicca (KCS). To assess the significance of laser confocal tomography of the cornea in the diagnosis and monitoring of KCS. We investigated 38 eyes of 30 patients with severe KCS. The patients were divided into two groups. Group 1 (15 patients, 19 eyes) was prescribed cyclosporine А 0.05% instillations 2 times daily, artificial tears, and soft contact lenses. Group 2 (15 patients, 19 eyes) received only instillations of cyclosporine А 0.05% 2 times daily and artificial tears. Besides standard ophthalmic examination, additional tests were performed, namely Schirmer's test, tear break-up time test, fluorescein eye stain test, tear osmolarity test (TearLab System, USA), and Heidelberg retinal tomography of the cornea (HRT, Heidelberg Engineering GmbH, Germany). HRT findings revealed a 3 times shorter epithelization period and faster recovery of corneal transparency in group 1 as compared to group 2 (1.5 and 4.5 months, respectively). There was also an evident reduction in the number of immune cells in the cornea, most pronounced in group 1 at 3 months, which is indicative of inflammation termination. The use of HRT of the cornea in KCS patients allows real-time cellular level observation of corneal changes, which together with clinical findings and diagnostic tests not only confirms the diagnosis but also determines treatment effectiveness. It has been also found that soft contact lenses accelerate epithelization of the cornea and relieve inflammation of the ocular surface in KCS patients under cyclosporine A 0.05% instillation therapy. Transparency of financial activity: the authors have no financial interest in the submitted materials and methods.

Zinc oxide tetrapods inhibit herpes simplex virus infection of cultured corneas

PubMed Central

Duggal, Neil; Jaishankar, Dinesh; Yadavalli, Tejabhiram; Hadigal, Satvik; Mishra, Yogendra Kumar; Adelung, Rainer

2017-01-01

Purpose Infection of the human cornea by herpes simplex virus type-1 (HSV-1) can cause significant vision loss. The purpose of this study was to develop an ex vivo model to visualize viral growth and spread in the cornea. The model was also used to analyze cytokine production and study the antiviral effects of zinc oxide tetrapods. Methods A β-galactosidase-expressing recombinant virus, HSV-1(KOS)tk12, was used to demonstrate the ability of the virus to enter and develop blue plaques on human corneal epithelial (HCE) cells and corneal tissues. Freshly obtained porcine corneas were cultured and then scratched before infection with HSV-1(KOS)tk12. The blue plaques on the corneas were imaged using a stereomicroscope. Western blot analysis for HSV-1 proteins was performed to verify HSV-1 infection of the cornea. Using the ex vivo model, zinc oxide tetrapods were tested for their anti-HSV-1 potential, and a cytokine profile was developed to assess the effects of the treatment. Results Cultured corneas and the use of β-galactosidase-expressing HSV-1(KOS)tk12 virus can provide an attractive ex vivo model to visualize and study HSV-1 entry and spread of the infection in tissues. We found that unlike cultured HCE cells, which demonstrated nearly 100% infectivity, HSV-1 infection of the cultured cornea was more restrictive and took longer to develop. We also found that the zinc oxide tetrapod–shaped nano- and microstructures inhibited HSV infection of the cultured cells, as well as the cultured corneas. The cytokine profile of the infected samples was consistent with previous studies of HSV-1 corneal infection. Conclusions The ability to visualize HSV-1 growth and spread in corneal tissues can provide new details about HSV-1 infection of the cornea and the efficacy of new cornea-specific antiviral drug candidates. The ex vivo model also demonstrates antiviral effects of zinc oxide tetrapods and adequately portrays the drug delivery issues that cornea-specific treatments face. PMID:28275313

A Simple and Reliable Technique to Orient Donor Corneal Tissue Using the Radial Width of the Surgical Limbus.

PubMed

Aldrich, Benjamin T; Stockman, Adam M; Freiburger, Monica J; Shinkunas, Todd J; Burckart, Kimberlee A; Schmidt, Gregory A; Reed, Cynthia R; Zimmerman, M Bridget; Goins, Kenneth M; Wagoner, Michael D; Greiner, Mark A

2015-12-01

To develop a method based on identification of the widest region of the surgical limbus that can yield quick and accurate orientation of excised human donor corneas. Corneoscleral tissue from donors 49 to 75 years old was marked at the temporal sclera at the time of recovery. Digital images obtained from 20 corneas stored in viewing chambers, retroilluminated and viewed from the endothelial side, were used to quantify the per-degree radial width of the surgical limbus, defined as the distance from the scleral spur to clear cornea. To evaluate differences in radial width among regions, measurements were compared with the intracorneal mean limbal width, and a per-degree z-score was calculated by averaging among corneas. Using images of corneas with the temporal mark masked and the sidedness known, 6 observers were subjected to a blinded trial of 10 corneas to determine the central point of the widest limbal region of each cornea. Compared with the intracorneal mean, the mean radial width of the surgical limbus was greatest in the superior quadrant, and the difference compared with the inferior, nasal, and temporal quadrants was significant (P < 0.0001). The superior region was identified with 100% accuracy in blinded trials. The average absolute difference between the predicted and actual central point of the superior limbus was 9.75 ± 0.30 degrees. The radial width of the surgical limbus is greatest in the superior region of the cornea and can be used as a diagnostic feature to orient donor corneal tissue.

Sorption of sodium hydroxide by type I collagen and bovine corneas.

PubMed

Whikehart, D R; Edwards, W C; Pfister, R R

1991-01-01

There are no quantitative studies on the uptake of alkali into corneal tissues. To study this phenomenon, both type I collagen and bovine corneas were incubated in sodium hydroxide (NaOH) under varying conditions for periods up to 27.5 h. The sorption (absorption or adsorption) of the alkali to protein and tissue was measured as the quantity of NaOH no longer available for titration to neutrality with hydrochloric acid. Sorption was found to be dependent on the concentration of NaOH (0.01-1 N) but independent of the incubation temperature (4-35 degrees C). In whole cornea, sorption of 1 N NaOH began immediately and increased with time up to 6 h. After 6 h, sorption decreased, together with the observed degradation and solubilization of the tissue. Stripping of the corneal endothelium alone or of the endothelium and epithelium increased sorption in a similar manner when compared to whole corneas for periods up to 4 h. These observations are compatible with ionic and nonionic bonding of hydroxide ions to collagen (including that of the cornea) and the subsequent release of hydroxide ions during hydrolysis of the protein itself. Indirect evidence also suggests the inclusion of quantities of unbound hydroxide ions in hydrated gels of glycosaminoglycans. It is proposed that in a chemical burn of the cornea, alkali is both stored in the tissue (by sorption) and reacted with it (by hydrolysis), without any net consumption of alkali taking place.

Validation of an endothelial roll preparation for Descemet Membrane Endothelial Keratoplasty by a cornea bank using "no touch" dissection technique.

PubMed

Marty, Anne-Sophie; Burillon, Carole; Desanlis, Adeline; Damour, Odile; Kocaba, Viridiana; Auxenfans, Céline

2016-06-01

Descemet Membrane Endothelial Keratoplasty (DMEK) selectively replaces the damaged posterior part of the cornea. However, the DMEK technique relies on a manually-performed dissection that is time-consuming, requires training and presents a potential risk of endothelial graft damages leading to surgery postponement when performed by surgeons in the operative room. To validate precut corneal tissue preparation for DMEK provided by a cornea bank in order to supply a quality and security precut endothelial tissue. The protocol was a technology transfer from the Netherlands Institute for Innovative Ocular Surgery (NIIOS) to Lyon Cornea Bank, after formation in NIIOS to the DMEK "no touch" dissection technique. The technique has been validated in selected conditions (materials, microscope) and after a learning curve, cornea bank technicians prepared endothelial tissue for DMEK. Endothelial cells densities (ECD) were evaluated before and after preparation, after storage and transport to the surgery room. Microbiological and histological controls have been done. Twenty corneas were manually dissected; 18 without tears. Nineteen endothelial grafts formed a double roll. The ECD loss after cutting was 3.3 % (n = 19). After transportation 7 days later, we found an ECD loss of 25 % (n = 12). Three days after cutting and transportation, we found 2.1 % of ECD loss (n = 7). Histology found an endothelial cells monolayer lying on Descemet membrane. The mean thickness was 12 ± 2.2 µm (n = 4). No microbial contamination was found (n = 19). Endothelial roll stability has been validated at 3 days in our cornea bank. Cornea bank technicians trained can deliver to surgeons an ECD controlled, safety and ready to use endothelial tissue, for DMEK by "no touch" technique, allowing time saving, quality and security for surgeons.

Comparison of Cornea Module and DermaInspect for noninvasive imaging of ocular surface pathologies

NASA Astrophysics Data System (ADS)

Steven, Philipp; Müller, Maya; Koop, Norbert; Rose, Christian; Hüttmann, Gereon

2009-11-01

Minimally invasive imaging of ocular surface pathologies aims at securing clinical diagnosis without actual tissue probing. For this matter, confocal microscopy (Cornea Module) is in daily use in ophthalmic practice. Multiphoton microscopy is a new optical technique that enables high-resolution imaging and functional analysis of living tissues based on tissue autofluorescence. This study was set up to compare the potential of a multiphoton microscope (DermaInspect) to the Cornea Module. Ocular surface pathologies such as pterygia, papillomae, and nevi were investigated in vivo using the Cornea Module and imaged immediately after excision by DermaInspect. Two excitation wavelengths, fluorescence lifetime imaging and second-harmonic generation (SHG), were used to discriminate different tissue structures. Images were compared with the histopathological assessment of the samples. At wavelengths of 730 nm, multiphoton microscopy exclusively revealed cellular structures. Collagen fibrils were specifically demonstrated by second-harmonic generation. Measurements of fluorescent lifetimes enabled the highly specific detection of goblet cells, erythrocytes, and nevus-cell clusters. At the settings used, DermaInspect reaches higher resolutions than the Cornea Module and obtains additional structural information. The parallel detection of multiphoton excited autofluorescence and confocal imaging could expand the possibilities of minimally invasive investigation of the ocular surface toward functional analysis at higher resolutions.

Engineering Functional Epithelium for Regenerative Medicine and In Vitro Organ Models: A Review

PubMed Central

Vrana, Nihal E.; Lavalle, Philippe; Dokmeci, Mehmet R.; Dehghani, Fariba; Ghaemmaghami, Amir M.

2013-01-01

Recent advances in the fields of microfabrication, biomaterials, and tissue engineering have provided new opportunities for developing biomimetic and functional tissues with potential applications in disease modeling, drug discovery, and replacing damaged tissues. An intact epithelium plays an indispensable role in the functionality of several organs such as the trachea, esophagus, and cornea. Furthermore, the integrity of the epithelial barrier and its degree of differentiation would define the level of success in tissue engineering of other organs such as the bladder and the skin. In this review, we focus on the challenges and requirements associated with engineering of epithelial layers in different tissues. Functional epithelial layers can be achieved by methods such as cell sheets, cell homing, and in situ epithelialization. However, for organs composed of several tissues, other important factors such as (1) in vivo epithelial cell migration, (2) multicell-type differentiation within the epithelium, and (3) epithelial cell interactions with the underlying mesenchymal cells should also be considered. Recent successful clinical trials in tissue engineering of the trachea have highlighted the importance of a functional epithelium for long-term success and survival of tissue replacements. Hence, using the trachea as a model tissue in clinical use, we describe the optimal structure of an artificial epithelium as well as challenges of obtaining a fully functional epithelium in macroscale. One of the possible remedies to address such challenges is the use of bottom-up fabrication methods to obtain a functional epithelium. Modular approaches for the generation of functional epithelial layers are reviewed and other emerging applications of microscale epithelial tissue models for studying epithelial/mesenchymal interactions in healthy and diseased (e.g., cancer) tissues are described. These models can elucidate the epithelial/mesenchymal tissue interactions at the microscale and provide the necessary tools for the next generation of multicellular engineered tissues and organ-on-a-chip systems. PMID:23705900

Engineering functional epithelium for regenerative medicine and in vitro organ models: a review.

PubMed

Vrana, Nihal E; Lavalle, Philippe; Dokmeci, Mehmet R; Dehghani, Fariba; Ghaemmaghami, Amir M; Khademhosseini, Ali

2013-12-01

Recent advances in the fields of microfabrication, biomaterials, and tissue engineering have provided new opportunities for developing biomimetic and functional tissues with potential applications in disease modeling, drug discovery, and replacing damaged tissues. An intact epithelium plays an indispensable role in the functionality of several organs such as the trachea, esophagus, and cornea. Furthermore, the integrity of the epithelial barrier and its degree of differentiation would define the level of success in tissue engineering of other organs such as the bladder and the skin. In this review, we focus on the challenges and requirements associated with engineering of epithelial layers in different tissues. Functional epithelial layers can be achieved by methods such as cell sheets, cell homing, and in situ epithelialization. However, for organs composed of several tissues, other important factors such as (1) in vivo epithelial cell migration, (2) multicell-type differentiation within the epithelium, and (3) epithelial cell interactions with the underlying mesenchymal cells should also be considered. Recent successful clinical trials in tissue engineering of the trachea have highlighted the importance of a functional epithelium for long-term success and survival of tissue replacements. Hence, using the trachea as a model tissue in clinical use, we describe the optimal structure of an artificial epithelium as well as challenges of obtaining a fully functional epithelium in macroscale. One of the possible remedies to address such challenges is the use of bottom-up fabrication methods to obtain a functional epithelium. Modular approaches for the generation of functional epithelial layers are reviewed and other emerging applications of microscale epithelial tissue models for studying epithelial/mesenchymal interactions in healthy and diseased (e.g., cancer) tissues are described. These models can elucidate the epithelial/mesenchymal tissue interactions at the microscale and provide the necessary tools for the next generation of multicellular engineered tissues and organ-on-a-chip systems.

Automated and Adaptable Quantification of Cellular Alignment from Microscopic Images for Tissue Engineering Applications

PubMed Central

Xu, Feng; Beyazoglu, Turker; Hefner, Evan; Gurkan, Umut Atakan

2011-01-01

Cellular alignment plays a critical role in functional, physical, and biological characteristics of many tissue types, such as muscle, tendon, nerve, and cornea. Current efforts toward regeneration of these tissues include replicating the cellular microenvironment by developing biomaterials that facilitate cellular alignment. To assess the functional effectiveness of the engineered microenvironments, one essential criterion is quantification of cellular alignment. Therefore, there is a need for rapid, accurate, and adaptable methodologies to quantify cellular alignment for tissue engineering applications. To address this need, we developed an automated method, binarization-based extraction of alignment score (BEAS), to determine cell orientation distribution in a wide variety of microscopic images. This method combines a sequenced application of median and band-pass filters, locally adaptive thresholding approaches and image processing techniques. Cellular alignment score is obtained by applying a robust scoring algorithm to the orientation distribution. We validated the BEAS method by comparing the results with the existing approaches reported in literature (i.e., manual, radial fast Fourier transform-radial sum, and gradient based approaches). Validation results indicated that the BEAS method resulted in statistically comparable alignment scores with the manual method (coefficient of determination R2=0.92). Therefore, the BEAS method introduced in this study could enable accurate, convenient, and adaptable evaluation of engineered tissue constructs and biomaterials in terms of cellular alignment and organization. PMID:21370940

Current status of synthetic epikeratoplasty.

PubMed

Thompson, K P; Hanna, K; Waring, G O; Gipson, I; Liu, Y; Gailitis, R P; Johnson-Wint, B; Green, K

1991-01-01

Many of the deficiencies with human tissue epikeratoplasty might be improved by the use of a suitable synthetic lenticule. Potential biomaterials for epikeratoplasty include collagen (types I, III, or IV), collagen-hydrogel copolymers, bioactive synthetics, and coated hydrogels. The biomaterial must be engineered to achieve strict specifications of optical clarity, support of epithelial migration and adhesion, permeability to solutes, and stability to corneal proteases. Attaching synthetic lenticules to the cornea without cutting Bowman's layer by adhesives, laser welding, or direct adhesion may also improve the efficacy of synthetic epikeratoplasty.

Transverse depth-dependent changes in corneal collagen lamellar orientation and distribution

PubMed Central

Abass, Ahmed; Hayes, Sally; White, Nick; Sorensen, Thomas; Meek, Keith M.

2015-01-01

It is thought that corneal surface topography may be stabilized by the angular orientation of out-of plane lamellae that insert into the anterior limiting membrane. In this study, micro-focus X-ray scattering data were used to obtain quantitative information about lamellar inclination (with respect to the corneal surface) and the X-ray scatter intensity throughout the depth of the cornea from the centre to the temporal limbus. The average collagen inclination remained predominantly parallel to the tissue surface at all depths. However, in the central cornea, the spread of inclination angles was greatest in the anterior-most stroma (reflecting the increased lamellar interweaving in this region), and decreased with tissue depth; in the peripheral cornea inclination angles showed less variation throughout the tissue thickness. Inclination angles in the deeper stroma were generally higher in the peripheral cornea, suggesting the presence of more interweaving in the posterior stroma away from the central cornea. An increase in collagen X-ray scatter was identified in a region extending from the sclera anteriorly until about 2 mm from the corneal centre. This could arise from the presence of larger diameter fibrils, probably of scleral origin, which are known to exist in this region. Incorporation of this quantitative information into finite-element models will further improve the accuracy with which they can predict the biomechanical response of the cornea to pathology and refractive procedures. PMID:25631562

Safety and Efficacy of OXB-202, a Genetically Engineered Tissue Therapy for the Prevention of Rejection in High-Risk Corneal Transplant Patients.

PubMed

Fouladi, Naghmeh; Parker, Maria; Kennedy, Vicky; Binley, Katie; McCloskey, Laura; Loader, Julie; Kelleher, Michelle; Mitrophanous, Kyriacos A; Stout, J Timothy; Ellis, Scott

2018-06-01

Due to both the avascularity of the cornea and the relatively immune-privileged status of the eye, corneal transplantation is one of the most successful clinical transplant procedures. However, in high-risk patients, which account for >20% of the 180,000 transplants carried out worldwide each year, the rejection rate is high due to vascularization of the recipient cornea. The main reason for graft failure is irreversible immunological rejection, and it is therefore unsurprising that neovascularization (NV; both pre and post grafting) is a significant risk factor for subsequent graft failure. NV is thus an attractive target to prevent corneal graft rejection. OXB-202 (previously known as EncorStat ® ) is a donor cornea modified prior to transplant by ex vivo genetic modification with genes encoding secretable forms of the angiostatic human proteins, endostatin and angiostatin. This is achieved using a lentiviral vector derived from the equine infectious anemia virus called pONYK1EiA, which subsequently prevents rejection by suppressing NV. Previously, it has been shown that rabbit donor corneas treated with pONYK1EiA substantially suppress corneal NV, opacity, and subsequent rejection in an aggressive rabbit model of cornea graft rejection. Here, efficacy data are presented in a second rabbit model, which more closely mirrors the clinical setting for high-risk corneal transplant patients, and safety data from a 3-month good laboratory practice toxicology and biodistribution study of pONYK1EiA-modified rabbit corneas in a rabbit corneal transplant model. It is shown that pONYK1EiA-modified rabbit corneas (OXB-202) significantly reduce corneal NV and the rate of corneal rejection in a dose-dependent fashion, and are tolerated with no adverse toxicological findings or significant biodistribution up to 13 weeks post surgery in these rabbit studies. In conclusion, angiogenesis is a valid target to prevent corneal graft rejection in a high-risk setting, and transplanted genetically modified corneas are safe and well-tolerated in an animal model. These data support the evaluation of OXB-202 in a first-in-human trial.

Changes of MK medium during storage of human cornea.

PubMed Central

Hasany, S M; Basu, P K

1987-01-01

By comparing the composition of McCarey-Kaufman (MK) medium before and after corneal storage we attempted to identify specific physiological changes in the medium as predictors of tissue damage. We also tried to determine if hydrocortisone (a lysosomal membrane stabiliser) added to the medium could reduce tissue damage during storage. Corneas (human and rabbit) were stored in the MK medium with and without hydrocortisone for 4 days at 4 degrees C. The water and nitrogen contents of the stored cornea were compared with those of the fresh cornea. The medium was analysed before and after corneal storage to determine the concentrations of glucose, protein, and amino acids as well as pH and osmolarity. Scanning electron microscopy (SEM) was used to estimate the degree of the corneal endothelial cell damage. The nitrogen contents and dry weights of the steroid treated and untreated stored corneas were similar to those of the fresh unstored cornea. The steroid treated cornea contained a lesser amount of water than the untreated cornea. The cornea stored in medium without steroid took up a greater amount of glucose from the medium than the cornea stored in medium with steroid. As compared with their concentrations in the fresh unused medium the concentrations of leucine, lysine, and glycine were lower and that of glutamic acid was higher in both the media used for corneal storage. However, the steroid treated storage medium as compared with the untreated storage medium had a greater reduction in the lowering of leucine, lysine, and glycine, and a lesser reduction in the increase of glutamic acid. Steroid treated medium also had a lesser amount of protein released from the stored cornea. Changes in the pH and osmolarity of the media before and after corneal storage were not remarkable. SEM showed that the endothelial cells of the cornea stored in the medium containing steroid were less damaged than those of the cornea stored in the medium without steroid. Images PMID:3620430

Using an ultrasound elasticity microscope to map three-dimensional strain in a porcine cornea.

PubMed

Hollman, Kyle W; Shtein, Roni M; Tripathy, Sakya; Kim, Kang

2013-08-01

An ultrasound elasticity microscope was used to map 3-D strain volume in an ex vivo porcine cornea to illustrate its ability to measure the mechanical properties of this tissue. Mechanical properties of the cornea play an important role in its function and, therefore, also in ophthalmic diseases such as kerataconus and corneal ectasia. The ultrasound elasticity microscope combines a tightly focused high-frequency transducer with confocal scanning to produce high-quality speckle over the entire volume of tissue. This system and the analysis were able to generate volume maps of compressional strain in all three directions for porcine corneal tissue, more information than any previous study has reported. Strain volume maps indicated features of the cornea and mechanical behavior as expected. These results constitute a step toward better understanding of corneal mechanics and better treatment of corneal diseases. Copyright © 2013 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.

In vivo study of the biocompatibility of a novel compressed collagen hydrogel scaffold for artificial corneas.

PubMed

Xiao, Xianghua; Pan, Shiyin; Liu, Xianning; Zhu, Xiuping; Connon, Che John; Wu, Jie; Mi, Shengli

2014-06-01

The experiments were designed to evaluate the biocompatibility of a plastically compressed collagen scaffold (PCCS). The ultrastructure of the PCCS was observed via scanning electron microscopy. Twenty New Zealand white rabbits were randomly divided into experimental and control groups that received corneal pocket transplantation with PCCS and an amniotic membrane, respectively. And the contralateral eye of the implanted rabbit served as the normal group. On the 1st, 7th, 14th, 21st, 30th, 60th, 90th, and 120th postoperative day, the eyes were observed via a slit lamp. On the 120th postoperative day, the rabbit eyes were enucleated to examine the tissue compatibility of the implanted stroma. The PCCS was white and translucent. The scanning electron microscopy results showed that fibers within the PCCS were densely packed and evenly arranged. No edema, inflammation, or neovascularization was observed on ocular surface under a slit lamp and few lymphocytes were observed in the stroma of rabbit cornea after histological study. In conclusion, the PCCS has extremely high biocompatibility and is a promising corneal scaffold for an artificial cornea. Copyright © 2013 Society of Plastics Engineers.

Atomic force microscopy analysis of human cornea surface after UV (λ=266 nm) laser irradiation

NASA Astrophysics Data System (ADS)

Spyratou, E.; Makropoulou, M.; Moutsouris, K.; Bacharis, C.; Serafetinides, A. A.

2009-07-01

Efficient cornea reshaping by laser irradiation for correcting refractive errors is still a major issue of interest and study. Although the excimer laser wavelength of 193 nm is generally recognized as successful in ablating corneal tissue for myopia correction, complications in excimer refractive surgery leads to alternative laser sources and methods for efficient cornea treatment. In this work, ablation experiments of human donor cornea flaps were conducted with the 4th harmonic of an Nd:YAG laser, with different laser pulses. AFM analysis was performed for examination of the ablated cornea flap morphology and surface roughness.

Preparation of pre-cut corneas from fresh donated whole globes for Descemet's stripping automated keratoplasty: 3-year results at the Central Eye Bank of Iran.

PubMed

Kanavi, Mozhgan Rezaei; Javadi, Mohammad Ali; Javadi, Fatemeh; Chamani, Tahereh

2014-09-01

To describe the technique and the results of the preparation of pre-cut corneas for Descemet's stripping automated endothelial keratoplasty (DSAEK) during a 3-year period at the Central Eye Bank of Iran (CEBI). The method of preparation of pre-cut corneas from donated whole globes at the CEBI is described and the frequency and percentage of pre-cut corneas prepared for DSAEK, between April 2009 and March 2012, are specified. Moreover, post-operative reports are reviewed for any complaints about using pre-cut tissues for DSAEK. Out of the 1,518 donated whole globes appropriate for DSAEK, 1,478 (97.4 %) pre-cut corneas were successfully prepared. The method of preparation failed in 40 (2.6 %) cases. Based on the eye bank post-operative reports, thickness of pre-cut tissues for DSAEK was deemed unacceptable in only 6 (0.4 %) cases prior to surgery; five of these were too thick and one was too thin. Preparation of pre-cut corneas, for DSAEK from donated whole globes, in the CEBI is a safe and easy method, with very good preservation of endothelial cells after the preparation of the pre-cut corneas and reduced risks from corneal manipulation.

Dendritic cells in the cornea during Herpes simplex viral infection and inflammation.

PubMed

Kwon, Min S; Carnt, Nicole A; Truong, Naomi R; Pattamatta, Ushasree; White, Andrew J; Samarawickrama, Chameen; Cunningham, Anthony L

Herpes simplex keratitis is commonly caused by Herpes simplex virus type 1, which primarily infects eyelids, corneas, or conjunctiva. Herpes simplex virus type 1-through sophisticated interactions with dendritic cells (DCs), a type of antigen-presenting cell)-initiates proinflammatory responses in the cornea. Corneas were once thought to be an immune-privileged region; however, with the recent discovery of DCs that reside in the cornea, this long-held conjecture has been overturned. Therefore, evaluating the clinical, preclinical, and cell-based studies that investigate the roles of DCs in corneas infected with Herpes simplex virus is critical. With in vivo confocal microscopy, animal models, and cell culture experiments, we may further the understanding of the sophisticated interactions of Herpes simplex virus with DCs in the cornea and the molecular mechanism associated with it. It has been shown that specific differentiation of DCs using immunohistochemistry, flow cytometry, and polymerase chain reaction analysis in both human and mice tissues and viral tissue infections are integral to increasing understanding. As for in vivo confocal microscopy, it holds promise as it is the least invasive and a real-time investigation. These tools will facilitate the discovery of various targets to develop new treatments. Copyright © 2017 Elsevier Inc. All rights reserved.

THz imaging system for in vivo human cornea.

PubMed

Sung, Shijun; Selvin, Skyler; Bajwa, Neha; Chantra, Somporn; Nowroozi, Bryan; Garritano, James; Goell, Jacob; Li, Alex; Deng, Sophie X; Brown, Elliott; Grundfest, Warren S; Taylor, Zachary D

2018-01-01

Terahertz (THz) imaging of corneal tissue water content (CTWC) is a proposed method for early, accurate detection and study of corneal diseases. Despite promising results from ex vivo and in vivo cornea studies, interpretation of the reflectivity data is confounded by the contact between corneal tissue and rigid dielectric window used to flatten the imaging field. This work develops a novel imaging system and image reconstruction methods specifically for nearly spherical targets such as human cornea. A prototype system was constructed using a 650 GHz multiplier source and Schottky diode detector. Resolution and imaging field strength measurement from characterization targets correlate well with those predicted by the quasioptical theory and physical optics analysis. Imaging experiments with corneal phantoms and ex vivo corneas demonstrate the hydration sensitivity of the imaging system and reliable measurement of CTWC. We present successful acquisition of non-contact THz images of in vivo human cornea, and discuss strategies for optimizing the imaging system design for clinical use.

Mitigating Scarring and Inflammation during Corneal Wound Healing using Nanofiber-Hydrogel Scaffolds

NASA Astrophysics Data System (ADS)

Fu, Amy

Due to the universal lack of donor tissue, there has been emerging interest in engineering materials to stimulate living cells to restore the features and functions of injured organs. We are particularly interested in developing materials for corneal use, where the necessity to maintain the tissue's transparency presents an additional challenge. Every year, there are 1.5 -- 2 million new cases of monocular blindness due to irregular healing of corneal injuries, dwarfing the approximately 150,000 corneal transplants performed. The large gap between the need and availability of cornea transplantation motivates us to develop a wound-healing scaffold that can prevent corneal blindness. To develop such a scaffold, it is necessary to regulate the cells responsible for repairing the damaged cornea, namely myofibroblasts, which are responsible for the disordered and non-refractive index matched scar that leads to corneal blindness. Using in vitro assays, we identified that protein nanofibers of certain orientation can promote cell migration and modulate the myofibroblast phenotype. The nanofibers are also transparent, easy to handle and non-cytotoxic. To adhere the nanofibers to a wound bed, we examined the use of two different in situ forming hydrogels: an artificial extracellular matrix protein (aECM)-based gel and a photo-crosslinkable heparin-based gel. Both hydrogels can be formed within minutes, are transparent upon gelation and are easily tunable. Using an in vivo mouse model for epithelial defects, we show that our corneal scaffolds (nanofibers together with hydrogel) are well-tolerated (no inflammatory response or turbidity) and support epithelium regrowth. We developed an ex vivo corneal tissue culture model where corneas that are wounded and treated with our scaffold can be cultured while retaining their ability to repair wounds for up to 21 days. Using this technique, we found that the aECM-based treatment induced a more favorable wound response than the heparin-based treatment, prompting us to further examine the efficacy of the aECM-based treatment in vivo using a rabbit model for stromal wounds. Results show that treated corneas have fewer myofibroblasts and immune cells than untreated ones, indicating that our corneal scaffold shows promise in promoting a calmer wound response and preventing corneal haze formation.

Corneal surface glycosylation is modulated by IL-1R and Pseudomonas aeruginosa challenge but is insufficient for inhibiting bacterial binding.

PubMed

Jolly, Amber L; Agarwal, Paresh; Metruccio, Matteo M E; Spiciarich, David R; Evans, David J; Bertozzi, Carolyn R; Fleiszig, Suzanne M J

2017-06-01

Cell surface glycosylation is thought to be involved in barrier function against microbes at mucosal surfaces. Previously we showed that the epithelium of healthy mouse corneas becomes vulnerable to Pseudomonas aeruginosa adhesion if it lacks the innate defense protein MyD88 (myeloid differentiation primary response gene 88), or after superficial injury by blotting with tissue paper. Here we explored their effect on corneal surface glycosylation using a metabolic label, tetra-acetylated N -azidoacetylgalactosamine (Ac 4 GalNAz). Ac 4 GalNAz treatment labeled the surface of healthy mouse corneas, leaving most cells viable, and bacteria preferentially associated with GalNAz-labeled regions. Surprisingly, corneas from MyD88 -/- mice displayed similar GalNAz labeling to wild-type corneas, but labeling was reduced and patchy on IL-1 receptor (IL-1R)-knockout mouse corneas ( P < 0.05, ANOVA). Tissue paper blotting removed GalNAz-labeled surface cells, causing DAPI labeling (permeabilization) of underlying cells. MS of material collected on the tissue paper blots revealed 67 GalNAz-labeled proteins, including intracellular proteins. These data show that the normal distribution of surface glycosylation requires IL-1R, but not MyD88, and is not sufficient to prevent bacterial binding. They also suggest increased P. aeruginosa adhesion to MyD88 -/- and blotted corneas is not due to reduction in total surface glycosylation, and for tissue paper blotting is likely due to cell permeabilization.-Jolly, A. L., Agarwal, P., Metruccio, M. M. E., Spiciarich, D. R., Evans, D. J., Bertozzi, C. R., Fleiszig, S. M. J. Corneal surface glycosylation is modulated by IL-1R and Pseudomonas aeruginosa challenge but is insufficient for inhibiting bacterial binding. © FASEB.

Corneal endothelium: developmental strategies for regeneration

PubMed Central

Zavala, J; López Jaime, G R; Rodríguez Barrientos, C A; Valdez-Garcia, J

2013-01-01

The main treatment available for restoration of the corneal endothelium is keratoplasty. This procedure is faced with several difficulties, including the shortage of donor tissue, post-surgical complications associated with the use of drugs to prevent immune rejection, and a significant increase in the occurrence of glaucoma. Recently, surgical procedures such as Descemet's stripping endothelial keratoplasty have focused on the transplant of corneal endothelium, yielding better visual results but still facing the need for donor tissue. The emergent strategies in the field of cell biology and tissue cultivation of corneal endothelial cells aim at the production of transplantable endothelial cell sheets. Cell therapy focuses on the culture of corneal endothelial cells retrieved from the donor, in the donor's cornea, followed by transplantation into the recipient. Recently, research has focused on overcoming the challenge of harvesting human corneal endothelial cells and the generation of new biomembranes to be used as cell scaffolds in surgical procedures. The use of corneal endothelial precursors from the peripheral cornea has also demonstrated to be effective and represents a valuable tool for reducing the risk of rejection in allogeneic transplants. Several animal model reports also support the use of adult stem cells as therapy for corneal diseases. Current results represent important progresses in the development of new strategies based on alternative sources of tissue for the treatment of corneal endotheliopathies. Different databases were used to search literature: PubMed, Google Books, MD Consult, Google Scholar, Gene Cards, and NCBI Books. The main search terms used were: ‘cornea AND embryology AND transcription factors', ‘human endothelial keratoplasty AND risk factors', ‘(cornea OR corneal) AND (endothelium OR endothelial) AND cell culture', ‘mesenchymal stem cells AND cell therapy', ‘mesenchymal stem cells AND cornea', and ‘stem cells AND (cornea OR corneal) AND (endothelial OR endothelium)'. PMID:23470788

A Mouse Model of the Cornea Pocket Assay for Angiogenesis Study

PubMed Central

Tang, Zhongshu; Zhang, Fan; Li, Yang; Arjunan, Pachiappan; Kumar, Anil; Lee, Chunsik; Li, Xuri

2011-01-01

A normal cornea is clear of vascular tissues. However, blood vessels can be induced to grow and survive in the cornea when potent angiogenic factors are administered 1. This uniqueness has made the cornea pocket assay one of the most used models for angiogenesis studies. The cornea composes multiple layers of cells. It is therefore possible to embed a pellet containing the angiogenic factor of interest in the cornea to investigate its angiogenic effect 2,3. Here, we provide a step by step demonstration of how to (I) produce the angiogenic factor-containing pellet (II) embed the pellet into the cornea (III) analyze the angiogenesis induced by the angiogenic factor of interest. Since the basic fibroblast growth factor (bFGF) is known as one of the most potent angiogenic factors 4, it is used here to induce angiogenesis in the cornea. PMID:21876523

Transient viscous response of the human cornea probed with the Surface Force Apparatus.

PubMed

Zappone, Bruno; Patil, Navinkumar J; Lombardo, Marco; Lombardo, Giuseppe

2018-01-01

Knowledge of the biomechanical properties of the human cornea is crucial for understanding the development of corneal diseases and impact of surgical treatments (e.g., corneal laser surgery, corneal cross-linking). Using a Surface Force Apparatus we investigated the transient viscous response of the anterior cornea from donor human eyes compressed between macroscopic crossed cylinders. Corneal biomechanics was analyzed using linear viscoelastic theory and interpreted in the framework of a biphasic model of soft hydrated porous tissues, including a significant contribution from the pressurization and viscous flow of fluid within the corneal tissue. Time-resolved measurements of tissue deformation and careful determination of the relaxation time provided an elastic modulus in the range between 0.17 and 1.43 MPa, and fluid permeability of the order of 10-13 m4/(N∙s). The permeability decreased as the deformation was increased above a strain level of about 10%, indicating that the interstitial space between fibrils of the corneal stromal matrix was reduced under the effect of strong compression. This effect may play a major role in determining the observed rate-dependent non-linear stress-strain response of the anterior cornea, which underlies the shape and optical properties of the tissue.

Impact of the Cornea Donor Study (CDS) on Acceptance of Corneas from Older Donors

PubMed Central

Sugar, Alan; Montoya, Monty M.; Beck, Roy; Cowden, John W.; Dontchev, Mariya; Gal, Robin L.; Kollman, Craig; Malling, Jackie; Mannis, Mark J.; Tennant, Bradley

2014-01-01

Purpose Evaluate retrospectively whether findings from the Cornea Donor Study (CDS) led to changes in the transplantation of corneas from older donors. Methods United States eye banks provided complete data on donor age and placement (domestic or international) for 86,273 corneas from 1998 to 2009. The data were analyzed by 3 time periods: preceding CDS (1998–1999), during CDS (2000–2007) and after publication of CDS 5 year results (2008–2009), and separately for corneas placed within vs. outside the United States. Results For corneal tissues transplanted in the United States, the percentage of donors ≥66 years old increased from 19% before CDS to 21% during CDS and 25% after CDS (p<0.001). Corresponding median (25th-75th percentile) donor ages were 53 (39–63), 54 (41–64) and 57 (46–66), respectively (p<0.001). The opposite trend was observed for corneas distributed outside the United States with the percentage of donors ≥66 years old decreasing from 56% to 42% to 34%, respectively. Donor age trends over time varied by eye bank. Conclusions There was a modest overall increase in the donor age of corneas transplanted in the United States from 1998 to 2009, but the retrospective nature of the study limits our ability to attribute this change to the CDS. The modest increases in the donor age of corneas transplanted is a positive finding, but wider acceptance of older corneal donor tissue should be encouraged based on the five-year evidence generated by the CDS. PMID:22262218

Raman Microscopy: A Noninvasive Method to Visualize the Localizations of Biomolecules in the Cornea.

PubMed

Kaji, Yuichi; Akiyama, Toshihiro; Segawa, Hiroki; Oshika, Tetsuro; Kano, Hideaki

2017-11-01

In vivo and in situ visualization of biomolecules without pretreatment will be important for diagnosis and treatment of ocular disorders in the future. Recently, multiphoton microscopy, based on the nonlinear interactions between molecules and photons, has been applied to reveal the localizations of various molecules in tissues. We aimed to use multimodal multiphoton microscopy to visualize the localizations of specific biomolecules in rat corneas. Multiphoton images of the corneas were obtained from nonlinear signals of coherent anti-Stokes Raman scattering, third-order sum frequency generation, and second-harmonic generation. The localizations of the adhesion complex-containing basement membrane and Bowman layer were clearly visible in the third-order sum frequency generation images. The fine structure of type I collagen was observed in the corneal stroma in the second-harmonic generation images. The localizations of lipids, proteins, and nucleic acids (DNA/RNA) was obtained in the coherent anti-Stokes Raman scattering images. Imaging technologies have progressed significantly and been applied in medical fields. Optical coherence tomography and confocal microscopy are widely used but do not provide information on the molecular structure of the cornea. By contrast, multiphoton microscopy provides information on the molecular structure of living tissues. Using this technique, we successfully visualized the localizations of various biomolecules including lipids, proteins, and nucleic acids in the cornea. We speculate that multiphoton microscopy will provide essential information on the physiological and pathological conditions of the cornea, as well as molecular localizations in tissues without pretreatment.

Elastic microfibril distribution in the cornea: Differences between normal and keratoconic stroma.

PubMed

White, Tomas L; Lewis, Philip N; Young, Robert D; Kitazawa, Koji; Inatomi, Tsutomu; Kinoshita, Shigeru; Meek, Keith M

2017-06-01

The optical and biomechanical properties of the cornea are largely governed by the collagen-rich stroma, a layer that represents approximately 90% of the total thickness. Within the stroma, the specific arrangement of superimposed lamellae provides the tissue with tensile strength, whilst the spatial arrangement of individual collagen fibrils within the lamellae confers transparency. In keratoconus, this precise stromal arrangement is lost, resulting in ectasia and visual impairment. In the normal cornea, we previously characterised the three-dimensional arrangement of an elastic fiber network spanning the posterior stroma from limbus-to-limbus. In the peripheral cornea/limbus there are elastin-containing sheets or broad fibers, most of which become microfibril bundles (MBs) with little or no elastin component when reaching the central cornea. The purpose of the current study was to compare this network with the elastic fiber distribution in post-surgical keratoconic corneal buttons, using serial block face scanning electron microscopy and transmission electron microscopy. We have demonstrated that the MB distribution is very different in keratoconus. MBs are absent from a region of stroma anterior to Descemet's membrane, an area that is densely populated in normal cornea, whilst being concentrated below the epithelium, an area in which they are absent in normal cornea. We contend that these latter microfibrils are produced as a biomechanical response to provide additional strength to the anterior stroma in order to prevent tissue rupture at the apex of the cone. A lack of MBs anterior to Descemet's membrane in keratoconus would alter the biomechanical properties of the tissue, potentially contributing to the pathogenesis of the disease. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Expression of pluripotency factors in larval epithelia of the frog Xenopus: Evidence for the presence of cornea epithelial stem cells

PubMed Central

Perry, Kimberly J.; Thomas, Alvin G.; Henry, Jonathan J.

2013-01-01

Understanding the biology of somatic stem cells in self renewing tissues represents an exciting field of study, especially given the potential to harness these cells for tissue regeneration and repair in treating injury and disease. The mammalian cornea contains a population of basal epithelial stem cells involved in cornea homeostasis and repair. Research has been restricted to mammalian systems and little is known about the presence or function of these stem cells in other vertebrates. Therefore, we carried out studies to characterize frog cornea epithelium. Careful examination shows that the Xenopus larval cornea epithelium consists of three distinct layers that include an outer epithelial layer and underlying basal epithelium, in addition to a deeper fibrous layer that contains the main sensory nerve trunks that give rise to numerous branches that extend into these epithelia. These nerves convey sensory and presumably also autonomic innervation to those tissues. The sensory nerves are all derived as branches of the trigeminal nerve/ganglion similar to the situation encountered in mammals, though there appear to be some potentially interesting differences, which are detailed in this paper. We show further that numerous pluripotency genes are expressed by cells in the cornea epithelium, including: sox2, p63, various oct4 homologs, c-myc, klf4 and many others. Antibody localization revealed that p63, a well known mammalian epithelial stem cell marker, was localized strictly to all cells in the basal cornea epithelium. c-myc, was visualized in a smaller subset of basal epithelial cells and adjacent stromal tissue predominately at the periphery of the cornea (limbal zone). Finally, sox2 protein was found to be present throughout all cells of both the outer and basal epithelia, but was much more intensely expressed in a distinct subset of cells that appeared to be either multinucleate or possessed multi-lobed nuclei that are normally located at the periphery of the cornea. Using a thymidine analog (EdU), we were able to label mitotically active cells, which revealed that cell proliferation takes place throughout the cornea epithelium, predominantly in the basal epithelial layer. Species of Xenopus and one other amphibian are unique in their ability to replace a missing lens from cells derived from the basal cornea epithelium. Using EdU we show, as others have previously, that proliferating cells within the cornea epithelium do contribute to the formation of these regenerated lenses. Furthermore, using qPCR we determined that representatives of various pluripotency genes (i.e., sox2, p63 and oct60) are upregulated early during the process of lens regeneration. Antibody labeling showed that the number of sox2 expressing cells increased dramatically within 4 hours following lens removal and these cells were scattered throughout the basal layer of the cornea epithelium. Historically, the process of lens regeneration in Xenopus had been described as one involving transdifferentiation of cornea epithelial cells (i.e., one involving cellular dedifferentiation followed by redifferentiation). Our combined observations provide evidence that a population of stem cells exists within the Xenopus cornea. We hypothesize that the basal epithelium contains oligopotent epithelial stem cells that also represent the source of regenerated lenses in the frog. Future studies will be required to clearly identify the source of these lenses. PMID:23274420

Multiparameter thermo-mechanical OCT-based characterization of laser-induced cornea reshaping

NASA Astrophysics Data System (ADS)

Zaitsev, Vladimir Yu.; Matveyev, Alexandr L.; Matveev, Lev A.; Gelikonov, Grigory V.; Vitkin, Alex; Omelchenko, Alexander I.; Baum, Olga I.; Shabanov, Dmitry V.; Sovetsky, Alexander A.; Sobol, Emil N.

2017-02-01

Phase-sensitive optical coherence tomography (OCT) is used for visualizing dynamic and cumulative strains and corneashape changes during laser-produced tissue heating. Such non-destructive (non-ablative) cornea reshaping can be used as a basis of emerging technologies of laser vision correction. In experiments with cartilaginous samples, polyacrilamide phantoms and excised rabbit eyes we demonstrate ability of the developed OCT system to simultaneously characterize transient and cumulated strain distributions, surface displacements, scattering tissue properties and possibility of temperature estimation via thermal-expansion measurements. The proposed approach can be implemented in perspective real-time OCT systems for ensuring safety of new methods of laser reshaping of cornea.

Genetics Home Reference: congenital stromal corneal dystrophy

MedlinePlus

... decorin, which is involved in the organization of collagens. Collagens are proteins that strengthen and support connective tissues ... ligaments. In the cornea, well-organized bundles of collagen make the cornea transparent. Decorin ensures that collagen ...

On the barrier properties of the cornea: a microscopy study of the penetration of fluorescently labeled nanoparticles, polymers, and sodium fluorescein.

PubMed

Mun, Ellina A; Morrison, Peter W J; Williams, Adrian C; Khutoryanskiy, Vitaliy V

2014-10-06

Overcoming the natural defensive barrier functions of the eye remains one of the greatest challenges of ocular drug delivery. Cornea is a chemical and mechanical barrier preventing the passage of any foreign bodies including drugs into the eye, but the factors limiting penetration of permeants and nanoparticulate drug delivery systems through the cornea are still not fully understood. In this study, we investigate these barrier properties of the cornea using thiolated and PEGylated (750 and 5000 Da) nanoparticles, sodium fluorescein, and two linear polymers (dextran and polyethylene glycol). Experiments used intact bovine cornea in addition to bovine cornea de-epithelialized or tissues pretreated with cyclodextrin. It was shown that corneal epithelium is the major barrier for permeation; pretreatment of the cornea with β-cyclodextrin provides higher permeation of low molecular weight compounds, such as sodium fluorescein, but does not enhance penetration of nanoparticles and larger molecules. Studying penetration of thiolated and PEGylated (750 and 5000 Da) nanoparticles into the de-epithelialized ocular tissue revealed that interactions between corneal surface and thiol groups of nanoparticles were more significant determinants of penetration than particle size (for the sizes used here). PEGylation with polyethylene glycol of a higher molecular weight (5000 Da) allows penetration of nanoparticles into the stroma, which proceeds gradually, after an initial 1 h lag phase.

Delivery of Riboflavin-5'-Monophosphate Into the Cornea: Can Liposomes Provide Any Enhancement Effects?

PubMed

Kandzija, Neva; Khutoryanskiy, Vitaliy V

2017-10-01

Keratoconus is a progressive condition caused by the thinning of the cornea, which eventually deforms the front surface of the eye into a cone shape leading to ghosting, multiple images, glare, and several other vision problems. Currently, keratoconus is treated with UV-induced riboflavin (Rb)-mediated collagen cross-linking, which requires a physical removal of the corneal epithelium under topical anesthesia. This study reports the penetration of Rb and its more water-soluble form, riboflavin-5'-monophosphate (RbP), into the bovine cornea ex vivo. Using ex vivo bovine corneal tissues and 0.8 mg/mL drug solutions in phosphate buffer, it was established that RbP penetration into the cornea within 3 h of diffusion experiment was greater (17.3 ± 0.8 μg) compared with Rb (10.4 ± 4.2 μg). In the cornea, RbP was found to convert to Rb, which is mediated with enzymes present in this tissue. Several formulations including the conventional and propylene glycol-containing liposomes with encapsulated RbP have been developed, and their effect on the drug penetration into the bovine cornea was evaluated. Encapsulation of RbP into the liposomes did not provide any statistically significant improvement in the penetration of RbP into the cornea. Copyright © 2017 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

Potentiation of femtosecond laser intratissue refractive index shaping (IRIS) in the living cornea with sodium fluorescein.

PubMed

Nagy, Lana J; Ding, Li; Xu, Lisen; Knox, Wayne H; Huxlin, Krystel R

2010-02-01

To assess the effectiveness of intratissue refractive index shaping (IRIS) in living corneas and test the hypothesis that it can be enhanced by increasing the two-photon absorption (TPA) of the tissue. Three corneas were removed from adult cats and cut into six pieces, which were placed in preservative (Optisol-GS; Bausch & Lomb, Inc., Irvine, CA) containing 0%, 0.25%, 1%, 1.5%, or 2.5% sodium fluorescein (Na-Fl). An 800-nm Ti:Sapphire femtosecond laser with a 100-fs pulse duration and 80-MHz repetition rate was used to perform IRIS in each piece, creating several refractive index (RI) modification lines at different speeds (between 0.1 and 5 mm/s). The lines were 1 mum wide, 10 microm apart, and approximately 150 microm below the tissue surface. The RI change of each grating was measured using calibrated, differential interference contrast microscopy. TUNEL staining was performed to assess whether IRIS or Na-Fl doping causes cell death. Scanning at 0.1 mm/s changed the RI of undoped, living corneas by 0.005. In doped corneas, RI changes between 0.01 and 0.02 were reliably achieved with higher scanning speeds. The magnitude of RI changes attained was directly proportional to Na-Fl doping concentration and inversely proportional to the scanning speed used to create the gratings. IRIS can be efficiently performed in living corneal tissue. Increasing the TPA of the tissue with Na-Fl increased both the scanning speeds and the magnitude of RI changes in a dose-dependent manner. Ongoing studies are exploring the use of IRIS to alter the optical properties of corneal tissue in situ, over an extended period.

Potentiation of Femtosecond Laser Intratissue Refractive Index Shaping (IRIS) in the Living Cornea with Sodium Fluorescein

PubMed Central

Nagy, Lana J.; Ding, Li; Xu, Lisen; Knox, Wayne H.

2010-01-01

Purpose. To assess the effectiveness of intratissue refractive index shaping (IRIS) in living corneas and test the hypothesis that it can be enhanced by increasing the two-photon absorption (TPA) of the tissue. Methods. Three corneas were removed from adult cats and cut into six pieces, which were placed in preservative (Optisol-GS; Bausch & Lomb, Inc., Irvine, CA) containing 0%, 0.25%, 1%, 1.5%, or 2.5% sodium fluorescein (Na-Fl). An 800-nm Ti:Sapphire femtosecond laser with a 100-fs pulse duration and 80-MHz repetition rate was used to perform IRIS in each piece, creating several refractive index (RI) modification lines at different speeds (between 0.1 and 5 mm/s). The lines were 1 μm wide, 10 μm apart, and ∼150 μm below the tissue surface. The RI change of each grating was measured using calibrated, differential interference contrast microscopy. TUNEL staining was performed to assess whether IRIS or Na-Fl doping causes cell death. Results. Scanning at 0.1 mm/s changed the RI of undoped, living corneas by 0.005. In doped corneas, RI changes between 0.01 and 0.02 were reliably achieved with higher scanning speeds. The magnitude of RI changes attained was directly proportional to Na-Fl doping concentration and inversely proportional to the scanning speed used to create the gratings. Conclusions. IRIS can be efficiently performed in living corneal tissue. Increasing the TPA of the tissue with Na-Fl increased both the scanning speeds and the magnitude of RI changes in a dose-dependent manner. Ongoing studies are exploring the use of IRIS to alter the optical properties of corneal tissue in situ, over an extended period. PMID:19815735

Tissue quality of eye-bank-prepared precut corneas for Descemet's stripping automated endothelial keratoplasty.

PubMed

Nelson, Brian A; Ritenour, Rusty J

2014-02-01

To evaluate endothelial cell density (ECD) of eye-bank-prepared tissue for use in Descemet's stripping automated endothelial keratoplasty (DSAEK). Prospective case series of consecutive corneal tissue prepared for DSAEK surgery. Sixty-seven sequential corneal-scleral tissue specimens representing 48 human donors processed for use in DSAEK surgery by the Regional Tissue Bank (Halifax, Nova Scotia). Corneal-scleral donor tissue was obtained by in situ recovery. ECD was recorded using the EB-3000 XYZ (HAI Laboratories Inc, Lexington, MA) specular microscope within 24 hours of preservation. Before the tissue was dissected, the corneal thickness was measured using the DGH-550 PACHETTE 2 (DGH Technology, Exton, PA) ultrasound pachymeter. The dissection was performed using a 300-μm Moria ALTK model microkeratome (Moria Inc). The posterior bed thickness was measured, and the anterior flap was replaced. Endothelial cell count density was obtained after re-preservation. Complete measurements were obtained for 42 of 67 corneas. In 25 corneas it was not possible to obtain a postdissection ECD measurement. The mean ECD before dissection was 2806 ± 317 cells/mm(2). The mean ECD after dissection was 2772 ± 318 cells/mm(2). There was an average loss of 34 cells/mm(2) (95% CI -110 to 40 cells/mm(2), p = 0.3). This case series confirms that ECD is preserved when DSAEK tissue is prepared in advance of surgery by trained eye-bank technicians in a low-volume Canadian eye bank. It was difficult to obtain clear images of the endothelial cell layer postdissection, possibly because of tissue swelling or distortion. Sixty-six of 67 corneas included in the study were used for surgery. © 2013 Canadian Ophthalmological Society Published by Canadian Ophthalmological Society All rights reserved.

Femtosecond laser cutting of human corneas for the subbasal nerve plexus evaluation.

PubMed

Kowtharapu, B S; Marfurt, C; Hovakimyan, M; Will, F; Richter, H; Wree, A; Stachs, O; Guthoff, R F

2017-01-01

Assessment of various morphological parameters of the corneal subbasal nerve plexus is a valuable method of documenting the structural and presumably functional integrity of the corneal innervation in health and disease. The aim of this work is to establish a rapid, reliable and reproducible method for visualization of the human corneal SBP using femtosecond laser cut corneal tissue sections. Trephined healthy corneal buttons were fixed and processed using TissueSurgeon-a femtosecond laser based microtome, to obtain thick tissue sections of the corneal epithelium and anterior stroma cut parallel to the ocular surface within approximately 15 min. A near infrared femtosecond laser was focused on to the cornea approximately 70-90 μm from the anterior surface to induce material separation using TissueSurgeon. The obtained corneal sections were stained following standard immunohistochemical procedures with anti-neuronal β-III tubulin antibody for visualization of the corneal nerves. Sections that contained the epithelium and approximately 20-30 μm of anterior stroma yielded excellent visualisation of the SBP with minimal optical interference from underlying stromal nerves. In conclusion, the results of this study have demonstrated that femtosecond laser cutting of the human cornea offers greater speed, ease and reliability than standard tissue preparation methods for obtaining high quality thick sections of the anterior cornea cut parallel to the ocular surface. © 2016 The Authors Journal of Microscopy © 2016 Royal Microscopical Society.

Design, clinical translation and immunological response of biomaterials in regenerative medicine

NASA Astrophysics Data System (ADS)

Sadtler, Kaitlyn; Singh, Anirudha; Wolf, Matthew T.; Wang, Xiaokun; Pardoll, Drew M.; Elisseeff, Jennifer H.

2016-07-01

The field of regenerative medicine aims to replace tissues lost as a consequence of disease, trauma or congenital abnormalities. Biomaterials serve as scaffolds for regenerative medicine to deliver cells, provide biological signals and physical support, and mobilize endogenous cells to repair tissues. Sophisticated chemistries are used to synthesize materials that mimic and modulate native tissue microenvironments, to replace form and to elucidate structure-function relationships of cell-material interactions. The therapeutic relevance of these biomaterial properties can only be studied after clinical translation, whereby key parameters for efficacy can be defined and then used for future design. In this Review, we present the development and translation of biomaterials for two tissue engineering targets, cartilage and cornea, both of which lack the ability to self-repair. Finally, looking to the future, we discuss the role of the immune system in regeneration and the potential for biomaterial scaffolds to modulate immune signalling to create a pro-regenerative environment.

Outcomes of descemet stripping automated endothelial keratoplasty using imported donor corneas.

PubMed

Lekhanont, Kaevalin; Vanikieti, Kavin; Nimvorapun, Nutthida; Chuckpaiwong, Varintorn

2017-04-05

The lack of development of local donor tissue acquisition in several regions of the world has resulted in the necessity of performing keratoplasty with imported donor corneas. The greatest concern about the use of donor corneas supplied by foreign eye banks is the effect of the increased donor death-to-operation time which inevitably occurs during the tissue recovery, tissue processing, and tissue transfer between the countries. The purpose of this study was to report the outcomes of descemet stripping automated endothelial keratoplasty (DSAEK) using imported donor corneas. This retrospective, non-comparative case series investigated the outcomes of the 102 consecutive DSAEK procedures using imported donor corneas performed at a single university-based hospital between August 2006-2014. The main outcome measures were postoperative best-corrected visual acuity (BCVA), endothelial cell density (ECD), and complications. The mean death-to-operation time was 9.52 ± 1.48 days (range, 8-13). The mean preoperative ECD was 2761 ± 285 cells/mm 2 . Fuchs' endothelial dystrophy was the predominant indication for grafting. The mean follow-up duration was 65.3 months. Ninety-three eyes had improved vision postoperatively (91.18%). BCVA unchanged in 3 eyes due to preexisting macular scar and advanced glaucoma. Primary graft failure occurred in 6 eyes (5.88%). Of the 93 eyes with improved BCVA, 100% had their best corrected vision within the first 1 year. The mean ECD at 6, 12, 24, 36, and 60 months after surgery was 1762 ± 294 cells/mm 2 , 1681 ± 284 cells/mm 2 , 1579 ± 209 cells/mm 2 , 1389 ± 273 cells/mm 2 , and 1251 ± 264 cells/mm 2 respectively. The mean ECD loss at 6 months, 1 year, 2 years, 3 years, and 5 years after surgery was 36.2%, 39.1%, 42.8%, 49.7%, and 54.7% respectively. The most common complication was graft detachment/dislocation (10.78%). There were no cases of any postoperative infection. DSAEK with imported donor corneas provides rapid and good visual rehabilitation. The percentages of endothelial cell loss were comparable to those achieved in Western series using domestic corneas in which fresher tissues were available for transplantation.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Akhtar, R.A.; Abdel-Latif, A.A.

Muscarinic cholinergic and ..cap alpha../sub 1/-adrenergic agonists provoke hydrolysis of PIP/sub 2/ into diacylglycerol (DG) and inositol trisphosphate (IP/sub 3/) in a wide variety of tissue. Recently, IP/sub 3/ has been shown to mobilize Ca/sup 2 +/ from ER in several permeabilized tissue preparations. Although rabbit cornea is enriched in ACh and NE, the physiological function of these neurotransmitters is unclear. The present studies were initiated to determine the effects of cholinergic and adrenergic agonists on PIP/sub 2/ turnover in the cornea. Addition of ACh or NE (50 ..mu..M each) to the /sup 32/P-labeled corneas for 10 min decreased themore » radioactivity in PIP/sub 2/ by 33 and 36%, and increased the radioactivity in phosphatidic acid by 72 and 52%, respectively. When the corneas were labeled with myo-(/sup 3/H)inositol, ACh and NE increased the accumulation of IP/sub 3/ by 92 and 48%, respectively. The effects of ACh and NE on phospholipid labeling and IP/sub 3/ accumulation were specifically inhibited by atropine (10 ..mu..M) and prazosin (10 ..mu..M), respectively. The data suggest the presence of muscarinic cholinergic and ..cap alpha../sub 1/-adrenergic receptors in the rabbit cornea. Furthermore, activation of these receptors leads to cleavage of PIP/sub 2/ into DG and IP/sub 3/ which may function as second messengers in this tissue.« less

3D Functional Corneal Stromal Tissue Equivalent Based on Corneal Stromal Stem Cells and Multi-Layered Silk Film Architecture.

PubMed

Ghezzi, Chiara E; Marelli, Benedetto; Omenetto, Fiorenzo G; Funderburgh, James L; Kaplan, David L

2017-01-01

The worldwide need for human cornea equivalents continues to grow. Few clinical options are limited to allogenic and synthetic material replacements. We hypothesized that tissue engineered human cornea systems based on mechanically robust, patterned, porous, thin, optically clear silk protein films, in combination with human corneal stromal stem cells (hCSSCs), would generate 3D functional corneal stroma tissue equivalents, in comparison to previously developed 2D approaches. Silk film contact guidance was used to control the alignment and distribution of hCSSCs on RGD-treated single porous silk films, which were then stacked in an orthogonally, multi-layered architecture and cultured for 9 weeks. These systems were compared similar systems generated with human corneal fibroblasts (hCFs). Both cell types were viable and preferentially aligned along the biomaterial patterns for up to 9 weeks in culture. H&E histological sections showed that the systems seeded with the hCSSCs displayed ECM production throughout the entire thickness of the constructs. In addition, the ECM proteins tested positive for keratocyte-specific tissue markers, including keratan sulfate, lumican, and keratocan. The quantification of hCSSC gene expression of keratocyte-tissue markers, including keratocan, lumican, human aldehyde dehydrogenase 3A1 (ALDH3A1), prostaglandin D2 synthase (PTDGS), and pyruvate dehydrogenase kinase, isozyme 4 (PDK4), within the 3D tissue systems demonstrated upregulation when compared to 2D single silk films and to the systems generated with the hCFs. Furthermore, the production of ECM from the hCSSC seeded systems and subsequent remodeling of the initial matrix significantly improved cohesiveness and mechanical performance of the constructs, while maintaining transparency after 9 weeks.

Intra-tissue Refractive Index Shaping (IRIS) of the cornea and lens using a low-pulse-energy femtosecond laser oscillator

PubMed Central

Ding, Li; Knox, Wayne H.; Bühren, Jens; Nagy, Lana J.; Huxlin, Krystel R.

2009-01-01

Purpose To assess the optical effect of high-repetition-rate, low energy femtosecond laser pulses on lightly-fixed corneas and lenses. Methods Eight corneas and eight lenses were extracted post-mortem from normal, adult cats. They were lightly fixed and stored in a solution that minimized swelling and opacification. An 800nm Ti:Sapphire femtosecond laser oscillator with a 27fs pulse duration and 93MHz repetition rate was used to inscribe gratings consisting of 20-40 lines, each 1μm wide, 100μm long and 5μm apart, 100μm below the tissue surface. Refractive index changes in the micromachined regions were calculated immediately and after one month of storage by measuring the intensity distribution of diffracted light when the gratings were irradiated with a 632.8nm He-Ne laser. Results Periodic gratings were created into the stromal layer of the corneas and the cortex of the lenses by adjusting the laser pulse energy until visible plasma luminescence and bubbles were no longer generated. The gratings had low scattering loss and could only be visualized using phase microscopy. Refractive index changes measured 0.005±0.001 to 0.01±0.001 in corneal tissue and 0.015±0.001 to 0.021±0.001 in the lenses. The gratings and refractive index changes were preserved after storing the micromachined corneas and lenses for one month. Conclusions These pilot experiments demonstrate a novel application of low-pulse-energy, MHz femtosecond lasers in modifying the refractive index of transparent ocular tissues without apparent tissue destruction. Although it remains to be verified in living tissues, the stability of this effect suggests that the observed modifications are due to long-term molecular and/or structural changes. PMID:18641284

Supercritical Carbon Dioxide-Assisted Decellularization of Aorta and Cornea.

PubMed

Guler, Selcan; Aslan, Bahar; Hosseinian, Pezhman; Aydin, Halil Murat

2017-09-01

Tissue engineering approaches utilize both natural and synthetic materials in the repair and regeneration processes. A naturally sourced material for this purpose is required to be free from any antigenic matter such as cells or cellular components. Decellularization of tissues may be achieved through chemical or physical removal agents. Supercritical carbon dioxide (sc-CO 2 ) has been used on the purpose of removing bioburden from tissues and offers an alternative to the traditionally used treatment methods. In addition to many advantages it offers with regard to the successful decellularization of tissues, it is known to have a sterilization effect. This study provides an insight into sc-CO 2 -assisted decellularization trials of corneal and aortic tissues. Results showed that high pressure of the fluid bursts the cells during the treatment and rapid depressurization was found to be effective in the removal of the cells from the tissues. sc-CO 2 decellularization offers significantly reduced treatment times, complete decellularization, and preserved extracellular matrix structure.

Finger's amniotic membrane buffer technique: protecting the cornea during radiation plaque therapy.

PubMed

Finger, Paul T

2008-04-01

To use amniotic membranes as a buffer between the cornea and radioactive eye plaques. Six melanomas were treated with ophthalmic plaque radiation therapy. Plaque-tumor localization required that a portion of the gold plaque touch the cornea during treatment. To enhance patient comfort and protect the cornea, an (0.1-mm-thick) amniotic membrane was interposed between the metal plaque edge and the cornea. Minimal ocular discomfort was noted during plaque radiation therapy. On a scale of 1 (none) to 10 (severe), all 6 patients reported pain levels of 1. As a tissue equivalent and because the mean thickness was only 0.1 mm, amniotic membranes had no significant effect on radiation dose calculations. No adverse effects, infections, or abrasions were noted. The amniotic membrane buffer technique improves patient comfort and protects the cornea during ophthalmic plaque radiation therapy.

EXPRESSION OF NeuGc ON PIG CORNEAS AND ITS POTENTIAL SIGNIFICANCE IN PIG CORNEAL XENOTRANSPLANTATION

PubMed Central

Lee, Whayoung; Miyagawa, Yuko; Long, Cassandra; Ekser, Burcin; Walters, Eric; Ramsoondar, Jagdeece; Ayares, David; Tector, A. Joseph; Cooper, David K. C.; Hara, Hidetaka

2016-01-01

Purpose Pigs expressing neither galactose-α1,3-galactose (Gal) nor N-glycolylneuraminic acid (NeuGc) take xenotransplantation one step closer to the clinic. Our aims were (i) to document the lack of NeuGc expression on corneas and aortas, and cultured endothelial cells (aortic [AECs]; corneal [CECs]) of GTKO/NeuGcKO pigs, and (ii) to investigate whether the absence of NeuGc reduced human antibody binding to the tissues and cells. Methods Wild-type (WT), GTKO, and GTKO/NeuGcKO pig were used for the study. Human tissues and cultured cells were negative controls. Immunofluorescence staining was performed using anti-Gal and anti-NeuGc antibodies, and to determine human IgM and IgG binding to tissues. Flow cytometric analysis was used to determine Gal and NeuGc expression on cultured CECs and AECs and to measure human IgM/IgG binding to these cells. Results Both Gal and NeuGc were detected on WT pig corneas and aortas. Although GTKO pigs expressed NeuGc, neither human nor GTKO/NeuGcKO pigs expressed Gal or NeuGc. Human IgM/IgG binding to corneas and aortas from GTKO and GTKO/NeuGcKO pigs was reduced compared to binding to WT pigs. Human antibody binding to GTKO/NeuGcKO AECs was significantly less than to GTKO AECs, but there was no significant difference in binding between GTKO and GTKO/NeuGcKO CECs. Conclusions The absence of NeuGc on GTKO aortic tissue and AECs is associated with reduced human antibody binding, and possibly will provide better outcome in clinical xenotransplantation using vascularized organs. For clinical corneal xenotransplantation, the absence of NeuGc expression on GTKO/NeuGcKO pig corneas may not prove an advantage over GTKO corneas. PMID:26418433

Co:MgF2 laser ablation of tissue: effect of wavelength on ablation threshold and thermal damage.

PubMed

Schomacker, K T; Domankevitz, Y; Flotte, T J; Deutsch, T F

1991-01-01

The wavelength dependence of the ablation threshold of a variety of tissues has been studied by using a tunable pulsed Co:MgF2 laser to determine how closely it tracks the optical absorption length of water. The Co:MgF2 laser was tuned between 1.81 and 2.14 microns, a wavelength region in which the absorption length varies by a decade. For soft tissues the ablation threshold tracks the optical absorption length; for bone there is little wavelength dependence, consistent with the low water content of bone. Thermal damage vs. wavelength was also studied for cornea and bone. Thermal damage to cornea has a weak wavelength dependence, while that to bone shows little wavelength dependence. Framing-camera pictures of the ablation of both cornea and liver show explosive removal of material, but differ as to the nature of the explosion.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Quantock,A.; Boote, C.; Young, R.

In the cornea of the eye light transmission is facilitated by the regular arrangement and uniform diameter of collagen fibrils that constitute the bulk of the extracellular corneal matrix. Matrix architecture, in turn, is believed to be governed by interactions between collagen fibrils and proteoglycan molecules modified with sulfated glycosaminoglycan side chains. Here, we outline the contribution made by small-angle X-ray scattering studies of the cornea in understanding the role of sulfated glycosaminoglycans in the control of collagen architecture in cornea, and present new depth-profiled microbeam data from swollen human eye-bank corneas that indicate no significant change in collagen fibrilmore » diameter throughout the tissue, but a lower collagen interfibrillar spacing in the anterior-most stromal regions compared with the ultrastructure of the deeper cornea.« less

In vitro 3D corneal tissue model with epithelium, stroma, and innervation.

PubMed

Wang, Siran; Ghezzi, Chiara E; Gomes, Rachel; Pollard, Rachel E; Funderburgh, James L; Kaplan, David L

2017-01-01

The interactions between corneal nerve, epithelium, and stroma are essential for maintaining a healthy cornea. Thus, corneal tissue models that more fully mimic the anatomy, mechanical properties and cellular components of corneal tissue would provide useful systems to study cellular interactions, corneal diseases and provide options for improved drug screening. Here a corneal tissue model was constructed to include the stroma, epithelium, and innervation. Thin silk protein film stacks served as the scaffolding to support the corneal epithelial and stromal layers, while a surrounding silk porous sponge supported neuronal growth. The neurons innervated the stromal and epithelial layers and improved function and viability of the tissues. An air-liquid interface environment of the corneal tissue was also mimicked in vitro, resulting in a positive impact on epithelial maturity. The inclusion of three cell types in co-culture at an air-liquid interface provides an important advance for the field of in vitro corneal tissue engineering, to permit improvements in the study of innervation and corneal tissue development, corneal disease, and tissue responses to environmental factors. Copyright © 2016 Elsevier Ltd. All rights reserved.

Contrasting cellular damage after Blue-IRIS and Femto-LASIK in cat cornea.

PubMed

Wozniak, Kaitlin T; Elkins, Noah; Brooks, Daniel R; Savage, Daniel E; MacRae, Scott; Ellis, Jonathan D; Knox, Wayne H; Huxlin, Krystel R

2017-12-01

Blue-intra-tissue refractive index shaping (Blue-IRIS) is a new approach to laser refractive correction of optical aberrations in the eye, which alters the refractive index of the cornea rather than changing its shape. Before it can be implemented in humans, it is critical to establish whether and to what extent, Blue-IRIS damages the cornea. Here, we contrasted the impact of -1.5 D cylinder refractive corrections inscribed using either Blue-IRIS or femtosecond laser in-situ keratomileusis (femto-LASIK) on corneal cell viability. Blue-IRIS was used to write a -1.5 D cylinder gradient index (GRIN) lens over a 2.5 mm by 2.5 mm area into the mid-stromal region of the cornea in six freshly-enucleated feline eyes. The same correction (-1.5 D cylinder) was inscribed into another four cat eyes using femto-LASIK. Six hours later, all corneas were processed for histology and stained for terminal deoxynucleotidyl transferase-mediated dUTP-digoxigenin nick end labeling (TUNEL) and p-γ-H2AX to label damaged cells. In Blue-IRIS-treated corneas, no tissue was removed and TUNEL-stained cells were confined to the laser focal zone in the stroma. In femto-LASIK, photoablation removed 14 μm of anterior stroma, but in addition, TUNEL-positive cells clustered across the femto-flap, the epithelium at the flap edges and the stroma below the ablation zone. Keratocytes positive for p-γ-H2AX were seen adjacent to all Blue-IRIS focal zones, but were completely absent from femto-LASIK-treated corneas. Unlike femto-LASIK, Blue-IRIS attains refractive correction in the cornea without tissue removal and only causes minimal, localized keratocyte death within the laser focal zones. In addition, Blue-IRIS induced DNA modifications associated with phosphorylation of γ-H2AX in keratocytes adjacent to the laser focal zones. We posit that this p-γ-H2AX response is related to alterations in chromatin structure caused by localized changes in osmolarity, a possible mechanism for the induced refractive index changes. Copyright © 2017 Elsevier Ltd. All rights reserved.

Cornea and ocular lens visualized with three-dimensional confocal microscopy

NASA Astrophysics Data System (ADS)

Masters, Barry R.

1992-08-01

This paper demonstrates the advantages of three-dimensional reconstruction of the cornea and the ocular crystalline lens by confocal microscopy and volume rendering computer techniques. The advantages of noninvasive observation of ocular structures in living, unstained, unfixed tissue include the following: the tissue is in a natural living state without the artifacts of fixation, mechanical sectioning, and staining; the three-dimensional structure can be observed from any view point and quantitatively analyzed; the dynamics of morphological changes can be studied; and the use of confocal microscopic observation results in a reduction of the number of animals required for ocular morphometric studies. The main advantage is that the dynamic morphology of ocular structures can be investigated in living ocular tissue. A laser scanning confocal microscope was used in the reflected light mode to obtain the two- dimensional images from the cornea and the ocular lens of a freshly enucleated rabbit eye. The light source was an argon ion laser with 488 nm wavelength. The microscope objective was a Leitz 25X, NA 0.6 water immersion lens. The 400 micron thick cornea was optically sectioned into 133, three micron sections. The semi-transparent cornea and the in-situ ocular lens was visualized as high resolution, high contrast two-dimensional images. The under sampling resulted in a three-dimensional visualization rendering in which the corneal thickness (z-axis) is compressed. The structures observed in the cornea include: superficial epithelial cells and their nuclei, basal epithelial cells and their `beaded' cell borders, basal lamina, nerve plexus, nerve fibers, free nerve endings in the basal epithelial cells, nuclei of stromal keratocytes, and endothelial cells. The structures observed in the in-situ ocular lens include: lens capsule, lens epithelial cells, and individual lens fibers.

[Cornea transplant].

PubMed

Garralda, A; Epelde, A; Iturralde, O; Compains, E; Maison, C; Altarriba, M; Goldaracena, M B; Maraví-Poma, E

2006-01-01

The keratoplasty, or cornea transplant, is one of the oldest surgical techniques in opthalmology, whose indication are: 1) tectonic, in order to preserve corneal anatomy and integrity; 2) clinical, in order to eliminate the inflamed corneal tissue in cases refractory to medical treatment; 3) optical, in order to improve visual acuity; and 4) cosmetic, in order to improve the appearance of the eye. Improvements in technique and instruments, as well as in post-operative treatment and the means of preserving donated tissue, have improved survival of the grafts. The Pamplona Model of transplant coordination of the Virgen del Camino Hospital is considered to be original and unique in Spain. The logistics of this program include the protocol for detection and extraction of corneas as well as for keratoplasties.

Cytochrome P450 Activity in Ex Vivo Cornea Models and a Human Cornea Construct.

PubMed

Kölln, Christian; Reichl, Stephan

2016-07-01

The pharmacokinetic behaviors of novel ophthalmic drugs are often preliminarily investigated in preclinical studies using ex vivo animal cornea or corneal cell culture models. During transcorneal passage, topically applied drugs may be affected by drug metabolizing enzymes. The knowledge regarding the functional expression of metabolic enzymes in corneal tissue is marginal; thus, the aim of this study was to investigate cytochrome P450 activity in an organotypic three-dimensional human cornea construct and to compare it with porcine and rabbit corneas, which are commonly used ex vivo cornea models. The total cytochrome P450 activity was determined by measuring the transformation of 7-ethoxycoumarin. Furthermore, the expression of the cytochrome P450 enzyme 2D6 (CYP2D6) was investigated at the protein level using immunohistochemistry and western blotting. CYP2D6 activity measurements were performed using a d-luciferin-based assay. In summary, similar levels of the total cytochrome P450 activity were identified in all 3 cornea models. The protein expression of CYP2D6 was confirmed in the human cornea construct and porcine cornea, whereas the signals in the rabbit cornea were weak. The analysis of the CYP2D6 activity indicated similar values for the human cornea construct and porcine cornea; however, a distinctly lower activity was observed in the rabbit cornea. Copyright © 2016 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

Effects of immersion solution on corneal tissue strip hydration and thickness during thermal shrinkage

NASA Astrophysics Data System (ADS)

Borja, David; Manns, Fabrice; Lamar, Peggy D.; Rosen, Alexander; Parel, Jean-Marie A.

2003-07-01

Purpose: To assess the effects of immersion solutions with different Dextran concentrations on the hydration of cornea tissue strips at normal body temperature. Methods: A 20% Dextran-BSS solution was injected via a self sealing limbal-transcorneal tunnel incision using a 30ga needle into the anterior chamber of human donor eyes until the globe was hard. The eyes were then immersed cornea down overnight in the same solution. Corneal thickness was measured by ultrasound pachymetry after the eyes were re-inflated and at regular intervals to assess dehydration. When the central cornea thickness reached 400-500μm corneal buttons were removed using a 10mm trephine. The buttons were then cut into 6x6mm strips using a custom-made jig and immediately immersed in solutions of Dextran (15 to 20% in increments of 2.5%) at 35degC. The edge thickness of the immersed strip was measured every 5 min for one hour using an optical comparator (Topcon, Japan) modified for tissue shadowphotogrammetry. Results: For five Florida Lions Eye Bank donated eyes after one hour in the Dextran solution the mean final measured thickness of corneas in 20%, 17.5% and 15% Dextran-BSS solutions were 570 (+/-75) μm, 680 (+/-70) μm, and 1080 (+/-95) μm respectively. These measured thicknesses changes correspond to an average swelling of 1.2, 1.4 and 2.2 times the initial thickness of each cornea strip in the 20%, 17.5% and 15% Dextran-BSS solutions respectively. Conclusion: We will conduct experiments at higher Dextran concentrations but our current results indicate that 20% Dextran-BSS solution was the best solution tested at keeping corneal tissue strips at or close to physiological thickness.

Ultrasonic biomicroscopy in ophthalmology and eye banking

NASA Astrophysics Data System (ADS)

Rosenwasser, George O. D.

1999-06-01

Echography has become a valuable diagnostic tool in ophthalmology. Ultrasonic biomicroscopy (UBM) in particular may be applied to the evaluation of small lesions of the anterior segment of the eye. Disease processes such as conjunctival and iris melanoma, other forms of neoplasia, intraocular cysts, narrow angle glaucoma, and intraocular foreign bodies can be diagnostically evaluated and followed longitudinally by UBM. Combining UBM with spectroscopy may become useful in determining cell type origins of a variety of tumors. Eye banking also has an increased need for UBM in corneal tissue banking. The recent development of the Laser In Situ Keratomileusis procedure has allowed corneal surgeries to create a partial thickness flap of tissue in the cornea, remove tissue from the base of the cornea with excimer laser ablation, and replace the hinged flap. This causes a substantial change in refractive error while thinning the cornea and leaving an interface within the corneal stroma. The ability to detect this type of surgery is essential in eye banking. Ultrasonic pachymetry to determine central thickness and biomicroscopy to detect the presence of an interface are essential in avoiding the use of these corneas for transplantation purposes. Determining the topography of the preserved corneas is another potential application for ultrasonography. Using this information to reduce optical aberration after transplant is crucial in improving visual performance post transplantation. A review of the anatomy of the eye, pathology of ocular diseases relevant to UBM, and principles of eye banking will be presented.

In-vitro corneal transparency measuring system

NASA Astrophysics Data System (ADS)

Ventura, Liliane; da Costa Vieira, Marcelo A.; Isaac, Flavio; Chiaradia, Caio; Faria de Sousa, Sidney J.

2001-06-01

A system for measuring the average corneal transparency of preserved corneas has been developed in order to provide a more accurate and standard report of the corneal tissue. The donated cornea transparency is one of the features to be analyzed previously to its indication for the transplant. The small portable system consists of two main parts: the optical and the electronic parts. The optical system consists of a white light, lenses and pin-holes that collimate white light beams that illuminates the cornea in its preservative medium. The light that passes through the cornea is detected by a resistive detector and the average corneal transparency is shown in a display. In order to obtain just the tissue transparency, the electronic circuit was built in a way that there is a baseline input of the preservative medium, previous to the measurement of the corneal transparency. Manipulating the system consists of three steps: (1) Adjusting the zero percentage in the absence of light (at this time the detectors in the dark); (2) Placing the preservative medium in the system and adjusting the 100% value (this is the baseline input); (3) Preserving the cornea and placing it in the system. The system provides the tissue transparency. The system is connected to an endothelium evaluation system for Slit Lamp, that we have developed, and statistics about the relationship of the corneal transparency and density of the endothelial cells will be provided in the next years. The system is being used in a public Eye Bank in Brasil.

A structural model for the in vivo human cornea including collagen-swelling interaction

PubMed Central

Cheng, Xi; Petsche, Steven J.; Pinsky, Peter M.

2015-01-01

A structural model of the in vivo cornea, which accounts for tissue swelling behaviour, for the three-dimensional organization of stromal fibres and for collagen-swelling interaction, is proposed. Modelled as a binary electrolyte gel in thermodynamic equilibrium, the stromal electrostatic free energy is based on the mean-field approximation. To account for active endothelial ionic transport in the in vivo cornea, which modulates osmotic pressure and hydration, stromal mobile ions are shown to satisfy a modified Boltzmann distribution. The elasticity of the stromal collagen network is modelled based on three-dimensional collagen orientation probability distributions for every point in the stroma obtained by synthesizing X-ray diffraction data for azimuthal angle distributions and second harmonic-generated image processing for inclination angle distributions. The model is implemented in a finite-element framework and employed to predict free and confined swelling of stroma in an ionic bath. For the in vivo cornea, the model is used to predict corneal swelling due to increasing intraocular pressure (IOP) and is adapted to model swelling in Fuchs' corneal dystrophy. The biomechanical response of the in vivo cornea to a typical LASIK surgery for myopia is analysed, including tissue fluid pressure and swelling responses. The model provides a new interpretation of the corneal active hydration control (pump-leak) mechanism based on osmotic pressure modulation. The results also illustrate the structural necessity of fibre inclination in stabilizing the corneal refractive surface with respect to changes in tissue hydration and IOP. PMID:26156299

Accelerated In Vitro Degradation of Optically Clear Low β-Sheet Silk Films by Enzyme-Mediated Pretreatment

PubMed Central

Shang, Ke; Rnjak-Kovacina, Jelena; Lin, Yinan; Hayden, Rebecca S.; Tao, Hu; Kaplan, David L.

2013-01-01

Purpose: To design patterned, transparent silk films with fast degradation rates for the purpose of tissue engineering corneal stroma. Methods: β-sheet (crystalline) content of silk films was decreased significantly by using a short water annealing time. Additionally, a protocol combining short water annealing time with enzymatic pretreatment of silk films with protease XIV was developed. Results: Low β-sheet content (17%–18%) and enzymatic pretreatment provided film stability in aqueous environments and accelerated degradation of the silk films in the presence of human corneal fibroblasts in vitro. The results demonstrate a direct relationship between reduced β-sheet content and enzymatic pretreatment, and overall degradation rate of the protein films. Conclusions: The novel protocol developed here provides new approaches to modulate the regeneration rate of silk biomaterials for corneal tissue regeneration needs. Translational Relevance: Patterned silk protein films possess desirable characteristics for corneal tissue engineering, including optical transparency, biocompatibility, cell alignment, and tunable mechanical properties, but current fabrication protocols do not provide adequate degradation rates to match the regeneration properties of the human cornea. This novel processing protocol makes silk films more suitable for the construction of human corneal stroma tissue and a promising way to tune silk film degradation properties to match corneal tissue regeneration. PMID:24049717

Accelerated in vitro Degradation of Optically Clear Low β-sheet Silk Films by Enzyme-Mediated Pretreatment

PubMed Central

Shang, Ke; Rnjak-Kovacina, Jelena; Lin, Yinan; Hayden, Rebecca S.; Hu, Tao; Kaplan, David L.

2013-01-01

Purpose To design patterned, transparent silk films with fast degradation rates for the purpose of tissue engineering corneal stroma, Methods β-sheet (crystalline) content of silk films was decreased significantly by using a short water annealing time. Additionally, a protocol combining short water annealing time with enzymatic pretreatment of silk films with protease XIV was developed. Results Low β-sheet content (17–18%) and enzymatic pre-treatment provided film stability in aqueous environments and accelerated degradation of the silk films in the presence of human corneal fibroblasts in vitro. The results demonstrate a direct relationship between reduced β-sheet content and enzymatic pre-treatment and overall degradation rate of the protein films. Conclusions The novel protocol developed here provides new approaches to modulate the regeneration rate of silk biomaterials for corneal tissue regeneration needs. Translational relevance Patterned silk protein films possess desirable characteristics for corneal tissue engineering, including optical transparency, biocompatibility, cell alignment and tunable mechanical properties, but current fabrication protocols do not provide adequate degradation rates to match the regeneration properties of the human cornea. This novel processing protocol makes silk films more suitable for the construction of human corneal stroma tissue and a promising way to tune silk film degradation properties to match corneal tissue regeneration. PMID:23579493

[The transfection and expression of IL-1ra gene to the rabbit cornea in situ via cation polymer mediation].

PubMed

Yuan, Jin; Chen, Jia-qi; Zhou, Shi-you; Liu, Zu-guo; Wang, Zhi-chong; Gu, Jian-jun

2006-08-01

To investigate the efficiency and safety of transfection of PEGFP-IL-1ra plasmid via cation polymer mediation (poly-ethylenimine, PEI) by injection into the corneal stroma. Human IL-1ra cDNA fragments were cloned by RT-PCR. Plasmid PEGFP-hIL-1ra recombinants were constructed and transferred into corneal endothelial cells (CEC) via cation polymer mediation. Expression of IL-1ra mRNA and IL-1ra was detected by green fluorescent protein (GFP) and Western-blotting. In the experiment group, 20 microl preparation containing 10 microg plasmid PEGFP-hIL-1ra recombinants and PEI-in-vivo was injected into the corneal stroma of Wistar rats (n = 30). Equivalent PEI-in-vivo solution was injected into another 15 corneas as the controls. Corneas were harvested at different time points (day 1, 3, 6, 14 and 21) after injection. The changes of tissue structure and function after IL-1ra in situ transfection were studied by HE staining, transmission electron microscopy, trypan blue-alizarin red staining and immunohistochemistry. The location and intensity of IL-1ra-GFP fusion protein expression were monitored by fluorescence microscopy. The size of the RT-PCR product of hIL-1ra fragments was approximately 500 bp in agarose gel electrophoresis. Restrictive enzyme digestion analysis of PstI, BamHI and DNA sequence analysis showed that expression of plasmid PEGFP-hIL-1ra recombinants had been constructed successfully. Twelve hours after the transfection of PEGFP-hIL-1ra, GFP fluorescence was detected in 10% - 15% endothelial cells. IL-1ra protein (RMW: 44,000) was detected by Western-blotting. In PEGFP-hIL-1ra treated group, fluorescence was appeared at day 1 in cornea basal epithelial cells, peaked at day 6 in whole cornea, began to weaken at day 14, and only weak fluorescence remained in cornea epithelial cells at day 21. No fluorescence appeared in the control group. No significant pathologic changes could be found in HE stained cornea tissues in both transfected group and the controls. p63 immunocytochemical staining in cornea epithelium was positive in both groups. Trypan blue-alizarin red staining confirmed that there was no damage in cornea endothelial cells. IL-1ra-GFP granules could be found by transmission electron microscope in every layer of cornea in the transfected group, but none in the controls. There was no impairment in the ultrastructure of cells in both groups. By direct injection of PEGFP-hIL-1ra into corneal stroma and mediated by cation polymer, IL-1ra genes could be transferred and expressed in corneal tissue efficiently and safely, and might provide a novel technique of gene transfection to cornea in situ.

[An experimental study of mesenchymal stem cells in tissue engineering scaffolds implanted in rabbit corneal lamellae to increase keratoprosthesis biointegration].

PubMed

Bai, H; Wang, L L; Huang, Y F; Huang, J X

2016-03-01

To complete a preliminary evaluation of the feasibility of implanting the complex of mouse bone marrow mesenchymal stem cells (BMSC) and a tissue engineering scaffold into rabbit corneal lamellae, based on which a solution may be proposed to consolidate the keratoprosthesis and the recipient surface, and to reduce the risk of complications. This experimental study was composed of two parts. (1) In vitro: some mouse BMSC were marked with red fluorescent proteins (RFP) and integrated with a decellularized pig articular cartilage extracellular matrix (ECM) scaffold. The cell survival was observed under a fluorescence microscope at 4 and 8 weeks. The cell distribution was examined by toluidine blue staining. The pore structure and the cell adhesion were observed under a scanning electron microscope. (2) in vivo: the complex of mouse BMSC and a decellularized scaffold was implanted into the lamellar cornea of 8 rabbit eyes with the fellow eyes as the controls. The eyes were sampled for observation using HE staining under a light microscope at 2, 4 and 8 weeks, respectively. The cell survival was examined under a fluorescence microscope, and the intracorneal cell survival at 8 weeks was observed using in vivo imaging. The conditions of ocular anterior segment of all the experimental animals were recorded. (1) Under the scanning electron microscope, the ECM scaffolds showed satisfactory porosity required for the adhesion and growth of cells and tissues, and the cell distribution over the cell-scaffold complex can be observed by toluidine blue staining. (2) Under the immunofluorescence microscope, cell proliferation was observed in vitro and in the interlamellar space (the maximum observation time was 8 weeks) after the RFP-marked mouse BMSC were integrated in vitro with ECM scaffolds. (3) Under the light microscope (HE staining), the stromal cells were detected to increase at each timepoint. A small number of monocytes and some mouse BMSC were observed in the superficial layer of corneal stroma, with sparsely and orderly arranged collagenous fibers and no neovascularization. All the epithelial cells appeared as mononuclear, columnar and undamaged, and the shape of ECM scaffolds, which were fused with the collagens, became unclear. (4) By in vivo imaging, it was found that the mouse BMSC survived for 8 weeks after being integrated with scaffolds and implanted into the interlamellar space of rabbit cornea. (5) After the implantation of cell-scaffold complex, severe postoperative inflammatory reactions, obvious conjunctival congestion and neovascularization were not observed. The corneal tissues surrounding the recipient area were transparent. One week later, mild inflammatory reactions were barely observed, and the cornea was transparent enough to observe the scaffold in the stromal layers. Four weeks later, the scaffolds became thinner. Eight weeks later, the scaffolds became extremely thin with normal vascular system in the corneal limbus. The ECM scaffold is a solid and biocompatible carrier for the growth and proliferation of BMSC. The mouse BMSC can grow and proliferate in the microenvironment of the interlamellar space of cornea.

Quantitative Analysis of Endothelial Cell Loss in Preloaded Descemet Membrane Endothelial Keratoplasty Grafts.

PubMed

Wolle, Meraf A; DeMill, David L; Johnson, Lauren; Lentz, Stephen I; Woodward, Maria A; Mian, Shahzad I

2017-11-01

Availability of preloaded Descemet membrane endothelial keratoplasty (pDMEK) tissue may increase acceptance of DMEK in surgical management of endothelial disease. The goal of this study was to determine the safety of pDMEK grafts for 24 hours before surgery by analyzing endothelial cell loss (ECL) using 2 image analysis software programs. A total of 18 cadaveric corneas were prepared for DMEK using a standardized technique and loaded in a modified Jones tube injector. Nine of the corneas were injected into Calcein AM vital dye after 1 minute (controls), and the remaining 9 corneas were left preloaded for 24 hours before injection into vital dye for staining. The stained corneas were imaged using an inverted confocal microscope. ECL was then analyzed and quantified by 2 different graders using 2 image analysis software programs. The control DMEK tissue resulted in 22.0% ± 4.0% ECL compared with pDMEK tissue, which resulted in 19.2% ± 7.2% ECL (P = 0.31). Interobserver agreement was 0.93 for MetaMorph and 0.92 for Fiji. The average time required to process images with MetaMorph was 2 ± 1 minutes and with Fiji was 20 ± 10 minutes. Intraobserver agreement was 0.97 for MetaMorph and 0.93 for Fiji. Preloading DMEK tissue is safe and may provide an alternative technique for tissue distribution and surgery for DMEK. The use of MetaMorph software for quantifying ECL is a novel and accurate imaging method with increased efficiency and reproducibility compared with the previously validated Fiji.

Application of erbium: YAG laser in ocular ablation.

PubMed

Tsubota, K

1990-01-01

Recent developments in lasers have provided us the possibility of laser ocular surgery. The xenon, argon, neodymium:YAG and dye lasers have been successfully used in out-patient clinics. The excimer laser has been attracting researchers' interest in the new application of laser to cornea and lens. The erbium:YAG laser emits a 2.94-microns beam that can ablate the transparent ocular tissues such as lenses and corneas. The author has applied this laser to the cornea, lens, vitreous and other ocular tissues. The erbium:YAG laser beam was directed through a 1.5-meter-long, 200-microns-diameter fiberoptic guide. The radiant energy measured about 50 mJ at the end of the probe. The laser was emitted as a 400-microsecond pulse. Freshly enucleated rabbit eyes were used in this study. Laser burns were applied to the tissue surface at various energy settings. At minimal power, the tissues were coagulated by the erbium:YAG laser application. At a power of more than 636-954 mJ/mm2, tissue began to evaporate; the tissue loss was observed under a surgical light microscope. Corneal photoablation, lens ablation, iridotomy, trabeculotomy, cutting of the vitreous and retinal ablation were easily performed. Like the excimer laser, the erbium:YAG laser is a potential tool for ocular surgery.

Assessment of corneal hydration sensing in the terahertz band: in vivo results at 100 GHz

NASA Astrophysics Data System (ADS)

Bennett, David; Taylor, Zachary; Tewari, Pria; Sung, Sijun; Maccabi, Ashkan; Singh, Rahul; Culjat, Martin; Grundfest, Warren; Hubschman, Jean-Pierre; Brown, Elliott

2012-09-01

Terahertz corneal hydration sensing has shown promise in ophthalmology applications and was recently shown to be capable of detecting water concentration changes of about two parts in a thousand in ex vivo corneal tissues. This technology may be effective in patient monitoring during refractive surgery and for early diagnosis and treatment monitoring in diseases of the cornea. In this work, Fuchs dystrophy, cornea transplant rejection, and keratoconus are discussed, and a hydration sensitivity of about one part in a hundred is predicted to be needed to successfully distinguish between diseased and healthy tissues in these applications. Stratified models of corneal tissue reflectivity are developed and validated using ex vivo spectroscopy of harvested porcine corneas that are hydrated using polyethylene glycol solutions. Simulation of the cornea's depth-dependent hydration profile, from 0.01 to 100 THz, identifies a peak in intrinsic reflectivity contrast for sensing at 100 GHz. A 100 GHz hydration sensing system is evaluated alongside the current standard ultrasound pachymetry technique to measure corneal hydration in vivo in four rabbits. A hydration sensitivity, of three parts per thousand or better, was measured in all four rabbits under study. This work presents the first in vivo demonstration of remote corneal hydration sensing.

Biointegration of the osteo-odonto lamina in the modified osteo-odonto keratoprosthesis: engineering of tissue to restore lost vision.

PubMed

Sawatari, Yoh; Marx, Robert E; Perez, Victor L; Parel, Jean-Marie

2013-01-01

The modified osteo-odonto keratoprosthesis (MOOKP) is a biologic keratoprosthesis that is used to treat a severely scarred cornea. The procedure involves multiple stages, including the transplantation of buccal mucosa to the damaged ocular surface and the implantation of an osteo-odonto lamina with a mounted polymethylmethacrylate lens. Among the keratoprostheses currently available, the MOOKP has proven to be the most effective based on the number of patients who have undergone the procedure and the duration of documented follow-up. Upon successful biointegration of the osteo-odonto lamina, the keratoprosthesis is able to resist resorption, provide stability, and prevent bacterial invasion and epithelial ingrowth. The effectiveness of the MOOKP is dependent on the anatomic and physiologic characteristics of the dental tissues and periodontal ligament.

Chemical Sympathectomy Increases Susceptibility to Ocular Herpes Simplex Virus Type 1 Infection

PubMed Central

Templeton, Amanda; Nguyen, Gabrielle; Ash, John D.; Straub, Rainer H.; Carr, Daniel J. J.

2008-01-01

The cornea is one of the most highly innervated tissues in the mammalian host. We hypothesized changes to cornea innervation through chemical sympathectomy would significantly alter the host response to the neurotropic viral pathogen, herpes simplex virus type 1 (HSV-1) following ocular infection. Mice treated with 6-hydroxydopamine hydrobromide displayed reduced tyrosine hydroxylase-positive fibers residing in the cornea. Sympathectomized mice were also found to show a transient rise in virus recovered in infected tissues and succumbed to infection in greater numbers. Whereas there were no differences in infiltrating leukocyte populations including HSV-1-specific cytotoxic T lymphocytes in the infected tissue, an increase in substance P and a decrease in IFN-γ levels in the trigeminal ganglion but not brain stem of sympathectomized mice were noted. Sympathectomized mice treated with the neurokinin-1 receptor antagonist L703,606 had delayed mortality implicating the involvement of substance P in HSV-1-mediated death. PMID:18495255

Measurement of an Elasticity Map in the Human Cornea

PubMed Central

Mikula, Eric R.; Jester, James V.; Juhasz, Tibor

2016-01-01

Purpose The biomechanical properties of the cornea have an important role in determining the shape of the cornea and visual acuity. Since the cornea is a nonhomogeneous tissue, it is thought that the elastic properties vary throughout the cornea. We aim to measure a map of corneal elasticity across the cornea. Methods An acoustic radiation force elasticity microscope (ARFEM) was used to create a map of corneal elasticity in the human cornea. This ARFEM uses a low frequency, high intensity acoustic force to displace a femtosecond laser-generated microbubble, while using a high frequency, low intensity ultrasound to monitor the position of the microbubble within the cornea. From the displacement of the bubble and the magnitude of the acoustic radiation force, the local value of corneal elasticity is calculated in the direction of the displacement. Measurements were conducted at 6 locations, ranging from the central to peripheral cornea at anterior and posterior depths. Results The mean anterior elastic moduli were 4.2 ± 1.2, 3.4 ± 0.7, and 1.9 ± 0.7 kPa in the central, mid, and peripheral regions, respectively, while the posterior elastic moduli were 2.3 ± 0.7, 1.6 ± 0.3, and 2.9 ± 1.2 kPa in the same radial locations. Conclusions We found that there is a unique distribution of elasticity axially and radially throughout the cornea. PMID:27327584

Modeling of ultrashort pulsed laser irradiation in the cornea based on parabolic and hyperbolic heat equations using electrical analogy

NASA Astrophysics Data System (ADS)

Gheitaghy, A. M.; Takabi, B.; Alizadeh, M.

2014-03-01

Hyperbolic and parabolic heat equations are formulated to study a nonperfused homogeneous transparent cornea irradiated by high power and ultrashort pulsed laser in the Laser Thermo Keratoplasty (LTK) surgery. Energy absorption inside the cornea is modeled using the Beer-Lambert law that is incorporated as an exponentially decaying heat source. The hyperbolic and parabolic bioheat models of the tissue were solved by exploiting the mathematical analogy between thermal and electrical systems, by using robust circuit simulation program called Hspice to get the solutions of simultaneous RLC and RC transmission line networks. This method can be used to rapidly calculate the temperature in laser-irradiated tissue at time and space domain. It is found that internal energy gained from the irradiated field results in a rapid rise of temperature in the cornea surface during the early heating period, while the hyperbolic wave model predicts a higher temperature rise than the classical heat diffusion model. In addition, this paper investigates and examines the effect of some critical parameters such as relaxation time, convection coefficient, radiation, tear evaporation and variable thermal conductivity of cornea. Accordingly, it is found that a better accordance between hyperbolic and parabolic models will be achieved by time.

Descemet membrane adhesion strength is greater in diabetics with advanced disease compared to healthy donor corneas.

PubMed

Schwarz, Chaid; Aldrich, Benjamin T; Burckart, Kimberlee A; Schmidt, Gregory A; Zimmerman, M Bridget; Reed, Cynthia R; Greiner, Mark A; Sander, Edward A

2016-12-01

Descemet membrane endothelial keratoplasty (DMEK) is an increasingly popular surgical procedure for treating ocular diseases that require a corneal transplant. Previous studies have found that tissue tearing during surgical preparation is more likely elevated in eyes from donors with a history of diabetes mellitus. To quantify these potential differences, we established an experimental technique for quantifying the force required to separate the endothelium-Descemet membrane complex (EDM) from stroma in human donor corneal tissue, and we assessed differences in adhesion strength between diabetic and non-diabetic donor corneas. Transplant suitable corneas were obtained from 23 donors 50-75 years old with an average preservation to assay time of 11.5 days. Corneas were classified from a medical records review as non-diabetic (ND, n = 9), diabetic without evidence of advanced disease (NAD, n = 8), or diabetic with evidence of advanced disease (AD, n = 10). Corneas were sectioned into 3 mm wide strips and the EDM peeled from the stroma. Using the force-extension data obtained from mechanical peel testing, EDM elastic peel tension (T E ), elastic stiffness (S E ), average delamination tension (T D ), and maximum tension (T MAX ) were calculated. Mean T E , S E , T D , and T MAX values for ND corneas were 0.78 ± 0.07 mN/mm, 0.37 ± 0.05 mN/mm/mm, 0.78 ± 0.08 mN/mm, and 0.94 ± 0.17 mN/mm, respectively. NAD values did not differ significantly. However, AD values for T E (1.01 ± 0.18 mN/mm), T D (1.09 ± 0.21 mN/mm), and T MAX (1.37 ± 0.24 mN/mm) were greater than ND and NAD corneas (P < 0.05). S E did not differ significantly between groups. These findings provide proof of the concept that chronic hyperglycemia from diabetes mellitus results in a phenotypically more adhesive interface between Descemet membrane and the posterior stroma in donor corneal tissue. Results of this study provide a foundation for further investigations into the impact of diabetes on the posterior cornea, eye banking, and keratoplasty. Copyright © 2016 Elsevier Ltd. All rights reserved.

Tissue procurement and transplantation: a Tuscany perspective.

PubMed

Filipponi, F; De Simone, P; Saviozzi, A; Bozzi, G

2008-01-01

Tissue procurement and transplantation are rarely taken into account as indicators of the efficiency of a regional donor procurement network. We present herein a retrospective review on Tuscany tissue procurement activities from 2004 until 2006. In 2003 the Tuscan Regional Government appointed a transplantation service authority to reorganize all regional donation and transplantation activities: the Organizzazione Toscana Trapianti (OTT). The regional tissue procurement network was based on either brain death (BD) and cardiac death (CD) donors under the responsibility of in-hospital transplantation coordinators (IHTCs). From 2004 to 2006, a total of 397 tissue donors were procured in Tuscany, and 4151 tissue transplantations were performed: 2909 skin grafts, 1209 bone grafts, and 33 heart valves. Over the same period, a total of 2116 cornea donors were procured; 4117 corneas were retrieved; 1779 were fit for transplantation, and 1418 were transplanted. Based on our experience, implementation of tissue procurement requires use of BD donors and paramount organizational efforts from IHTCs.

Selective two-photon collagen crosslinking in situ measured by Brillouin microscopy (Conference Presentation)

NASA Astrophysics Data System (ADS)

Kwok, Sheldon J. J.; Kuznetsov, Ivan A.; Kim, Moonseok; Choi, Myunghwan; Scarcelli, Giuliano; Yun, Seok-Hyun

2017-02-01

Two-photon polymerization and crosslinking are commonly used methods for microfabrication of three-dimensional structures with applications spanning from photonic microdevices, drug delivery systems, to cellular scaffolds. However, the use of two-photon processes for precise, internal modification of biological tissues has not yet been reported. One of the major challenges has been a lack of appropriate tools to monitor and characterize crosslinked regions nondestructively. Here, we demonstrate spatially selective two-photon collagen crosslinking (2P-CXL) in intact tissue for the first time. Using riboflavin photosensitizer and femtosecond laser irradiation, we crosslinked a small volume of tissue within animal corneas. Collagen fiber orientations and photobleaching were characterized by second harmonic generation and two-photon fluorescence imaging, respectively. Using confocal Brillouin microscopy, we measured local changes in longitudinal mechanical moduli and visualized the cross-linked pattern without perturbing surrounding non-irradiated regions. 2P-CXL-induced tissue stiffening was comparable to that achieved with conventional one-photon CXL. Our results demonstrate the ability to selectively stiffen biological tissue in situ at high spatial resolution, with broad implications in ophthalmology, laser surgery, and tissue engineering.

The Ets Transcription Factor EHF as a Regulator of Cornea Epithelial Cell Identity*

PubMed Central

Stephens, Denise N.; Klein, Rachel Herndon; Salmans, Michael L.; Gordon, William; Ho, Hsiang; Andersen, Bogi

2013-01-01

The cornea is the clear, outermost portion of the eye composed of three layers: an epithelium that provides a protective barrier while allowing transmission of light into the eye, a collagen-rich stroma, and an endothelium monolayer. How cornea development and aging is controlled is poorly understood. Here we characterize the mouse cornea transcriptome from early embryogenesis through aging and compare it with transcriptomes of other epithelial tissues, identifying cornea-enriched genes, pathways, and transcriptional regulators. Additionally, we profiled cornea epithelium and stroma, defining genes enriched in these layers. Over 10,000 genes are differentially regulated in the mouse cornea across the time course, showing dynamic expression during development and modest expression changes in fewer genes during aging. A striking transition time point for gene expression between postnatal days 14 and 28 corresponds with completion of cornea development at the transcriptional level. Clustering classifies co-expressed, and potentially co-regulated, genes into biologically informative categories, including groups that exhibit epithelial or stromal enriched expression. Based on these findings, and through loss of function studies and ChIP-seq, we show that the Ets transcription factor EHF promotes cornea epithelial fate through complementary gene activating and repressing activities. Furthermore, we identify potential interactions between EHF, KLF4, and KLF5 in promoting cornea epithelial differentiation. These data provide insights into the mechanisms underlying epithelial development and aging, identifying EHF as a regulator of cornea epithelial identity and pointing to interactions between Ets and KLF factors in promoting epithelial fate. Furthermore, this comprehensive gene expression data set for the cornea is a powerful tool for discovery of novel cornea regulators and pathways. PMID:24142692

From Nano to Macro: Studying the Hierarchical Structure of the Corneal Extracellular Matrix

PubMed Central

Quantock, Andrew J.; Winkler, Moritz; Parfitt, Geraint J.; Young, Robert D.; Brown, Donald J.; Boote, Craig; Jester, James V.

2014-01-01

In this review, we discuss current methods for studying ocular extracellular matrix (ECM) assembly from the ‘nano’ to the ‘macro’ levels of hierarchical organization. Since collagen is the major structural protein in the eye, providing mechanical strength and controlling ocular shape, the methods presented focus on understanding the molecular assembly of collagen at the nanometer level using x-ray scattering through to the millimeter to centimeter level using nonlinear optical (NLO) imaging of second harmonic generated (SHG) signals. Three-dimensional analysis of ECM structure is also discussed, including electron tomography, serial block face scanning electron microscopy (SBF-SEM) and digital image reconstruction. Techniques to detect non-collagenous structural components of the ECM are also presented, and these include immunoelectron microscopy and staining with cationic dyes. Together, these various approaches are providing new insights into the structural blueprint of the ocular ECM, and in particular that of the cornea, which impacts upon our current understanding of the control of corneal shape, pathogenic mechanisms underlying ectatic disorders of the cornea and the potential for corneal tissue engineering. PMID:25819457

Laser welding of biological tissue: experimental studies in ophthalmology

NASA Astrophysics Data System (ADS)

Pini, Roberto; Rossi, Francesca; Menabuoni, Luca

2006-04-01

In this paper we present an original approach to laser welding of ocular media. Attention is focused on laser welding of the cornea and lens capsule. The process is based on the interaction of near infrared diode laser radiation (at 810 nm) with tissue that was previously stained with an Indocyanine Green solution in sterile water. The topical application of the chromophore makes possible a selective heating of the tissue, which results in a homogenous welding effect with low thermal damage to the surrounding tissue. Experimental tests were performed ex vivo on both capsule and cornea, and in vivo (rabbits) only on the cornea, in order to characterize the process as a whole. Spectrophotometric, biomechanical, and thermal measurements were carried out in order to study the laser-tissue interaction, while morphological, histological and auto-florescence microscopy analyses made during a follow-up study provided information on the healing process in welded rabbit corneas. The welding procedure was set up according to the type of tissue, with the staining procedure and irradiation conditions being optimized in each case. Our test indicated that: 1) laser welding of corneal wounds, which is a non contact technique performed at low continuous wave laser power (12 W/cm2), can be proposed as a support to or substitute for the standard suturing technique in cataract surgery and in penetrating keratoplasty (in corneal transplants); 2) laser welding of the lens capsule requires a "contact irradiating technique" in order to be efficiently performed, since the tissue is in underwater conditions, with single spot pulses of about 100 J/cm2 fluence and pulse duration around 100 ms. In the latter case, laser welding was proposed as a tool for assisting closure of the lens capsule after the lens refilling procedure (Phaco-ersatz), or for repairing capsular breaks induced by accidental traumas or produced intraoperatively.

Evaluation of prior photorefractive keratectomy in donor tissue.

PubMed

Terry, M A; Ousley, P J; Rich, L F; Wilson, D J

1999-05-01

To describe a case in which an eye donor had prior bilateral photorefractive keratectomies and to elucidate possible methods of evaluation and screening of donor tissue. Case report. A 62-year-old eye donor was reported to have received radial keratotomy before his death. Further investigation by the eye bank showed a history of photorefractive keratectomy (PRK), not radial keratotomy. The corneas were therefore not used for transplantation, and the eyes were evaluated by slit-lamp examination, photography, corneal topography, and histology. Slit-lamp and photographic examination did not indicate the presence of PRK ablations. Corneal topography mapping with the TMS-1 was relatively ambiguous for identifying PRK flattening, while multiple data formatting of the cornea with the Orbscan resulted in the strongest suggestion of prior PRK. Histologic analysis showed central corneal thinning and loss of Bowman's membrane consistent with PRK. In the absence of a positive donor history for PRK, current methods of screening donor tissue for prior PRK often are insufficient to exclude these corneas from use in transplantation. More refined placido imagery corneal topography or newer technologies such as the Orbscan may allow more sensitive and specific methods of donor tissue screening.

[Comparative analysis of the donor cornea quality and of the interval between death and preservation before and after new sanitary and technique rules in a University Eye Bank].

PubMed

Zantut, Fabio; Holzchuh, Ricardo; Boni, Reginaldo Carlos; Mackus, Eva Cristina; Zantut, Paulo Roberto; Nakano, Claudio; Netto, Adamo Lui; Hida, Richard Yudi

2012-01-01

To compare the interval between death and enucleation (ΔT-O-E), between enucleation and preservation (ΔT-E-P) and the quality of the cornea before and after the implantation of new technique and sanitary rules. A retrospective study that evaluated the records of cornea donors in Sao Paulo's Santa Casa Eye Tissue Bank 2 years before and 2 years after the implementation new sanitary rules. An increase was observed in the absolute number of 205 to 374 donors following the adopted changes. There was no statistically significant difference in Δt-O-E and ΔT-E-P before and after the implemented changes. Of the total of 1,105 donor corneas, 388 donor corneas were observed before the changes and 717 donor corneas after the implemented changes. We observed a statistically significant increase in grading of donor cornea quality from 1.76 ± 0.90 to 1.94 ± 0.88 after the implementation of new standards of resolution. After the changes required by Resolution 347, there was a large increase in the number of donated, taken and preserved corneas. The BTO has not diminished the ΔT O-E and ΔT E-P. Cornea quality presented itself lower after the new rules.

The influence of intraocular pressure and air jet pressure on corneal contactless tonometry tests.

PubMed

Simonini, Irene; Pandolfi, Anna

2016-05-01

The air puff is a dynamic contactless tonometer test used in ophthalmology clinical practice to assess the biomechanical properties of the human cornea and the intraocular pressure due to the filling fluids of the eye. The test is controversial, since the dynamic response of the cornea is governed by the interaction of several factors which cannot be discerned within a single measurement. In this study we describe a numerical model of the air puff tests, and perform a parametric analysis on the major action parameters (jet pressure and intraocular pressure) to assess their relevance on the mechanical response of a patient-specific cornea. The particular cornea considered here has been treated with laser reprofiling to correct myopia, and the parametric study has been conducted on both the preoperative and postoperative geometries. The material properties of the cornea have been obtained by means of an identification procedure that compares the static biomechanical response of preoperative and postoperative corneas under the physiological IOP. The parametric study on the intraocular pressure suggests that the displacement of the cornea׳s apex can be a reliable indicator for tonometry, and the one on the air jet pressure predicts the outcomes of two or more distinct measurements on the same cornea, which can be used in inverse procedures to estimate the material properties of the tissue. Copyright © 2015 Elsevier Ltd. All rights reserved.

Silk fibroin/chitosan thin film promotes osteogenic and adipogenic differentiation of rat bone marrow-derived mesenchymal stem cells.

PubMed

Li, Da-Wei; He, Jin; He, Feng-Li; Liu, Ya-Li; Liu, Yang-Yang; Ye, Ya-Jing; Deng, Xudong; Yin, Da-Chuan

2018-04-01

As a biodegradable polymer thin film, silk fibroin/chitosan composite film overcomes the defects of pure silk fibroin and chitosan films, respectively, and shows remarkable biocompatibility, appropriate hydrophilicity and mechanical properties. Silk fibroin/chitosan thin film can be used not only as metal implant coating for bone injury repair, but also as tissue engineering scaffold for skin, cornea, adipose, and other soft tissue injury repair. However, the biocompatibility of silk fibroin/chitosan thin film for mesenchymal stem cells, a kind of important seed cell of tissue engineering and regenerative medicine, is rarely reported. In this study, silk fibroin/chitosan film was prepared by solvent casting method, and the rat bone marrow-derived mesenchymal stem cells were cultured on the silk fibroin/chitosan thin film. Osteogenic and adipogenic differentiation of rat bone marrow-derived mesenchymal stem cells were induced, respectively. The proliferation ability, osteogenic and adipogenic differentiation abilities of rat bone marrow-derived mesenchymal stem cells were systematically compared between silk fibroin/chitosan thin film and polystyrene tissue culture plates. The results showed that silk fibroin/chitosan thin film not only provided a comparable environment for the growth and proliferation of rat bone marrow-derived mesenchymal stem cells but also promoted their osteogenic and adipogenic differentiation. This work provided information of rat bone marrow-derived mesenchymal stem cells behavior on silk fibroin/chitosan thin film and extended the application of silk fibroin/chitosan thin film. Based on the results, we suggested that the silk fibroin/chitosan thin film could be a promising material for tissue engineering of bone, cartilage, adipose, and skin.

Nonlinear side effects of fs pulses inside corneal tissue during photodisruption

NASA Astrophysics Data System (ADS)

Heisterkamp, A.; Ripken, T.; Mamom, T.; Drommer, W.; Welling, H.; Ertmer, W.; Lubatschowski, H.

In order to evaluate the potential for refractive surgery, fs laser pulses of 150-fs pulse duration were used to process corneal tissue of dead and living animal eyes. By focusing the laser radiation down to spot sizes of several microns, very precise cuts could be achieved inside the treated cornea, accompanied with minimum collateral damage to the tissue by thermal or mechanical effects. During histo-pathological analysis by light and transmission electron microscopy considerable side effects of fs photodisruption were found. Due to the high intensities at the focal region several nonlinear effects occurred. Self-focusing, photodissociation, UV-light production were observed, leading to streak formation inside the cornea.

Mass Spectrometric Analyses of Phosphatidylcholines in Alkali-Exposed Corneal Tissue

PubMed Central

Crane, Ashley M.; Hua, Hong-Uyen; Coggin, Andrew D.; Gugiu, Bogdan G.; Lam, Byron L.; Bhattacharya, Sanjoy K.

2012-01-01

Purpose. The aims were to determine whether exposure to sodium hydroxide results in predictable changes in phosphatidylcholine (PC) in corneal tissue and if PC profile changes correlate to exposure duration. PCs are major components of the cell membrane lipid bilayer and are often involved in biological processes such as signaling. Methods. Enucleated porcine (n = 140) and cadaver human eyes (n = 20) were exposed to water (control) and 11 M NaOH. The corneas were excised and lipids were extracted using the Bligh and Dyer method with suitable modifications. Class-specific lipid identification was carried out using a ratiometric lipid standard on a TSQ Quantum Access Max mass spectrometer. Protein amounts were determined using Bradford assays. Results. Control and alkali-treated corneas showed reproducible PC spectra for both porcine and human corneas. Over 200 PCs were identified for human and porcine control and each experimental time point. Several PC species (m/z values) consequent upon alkali exposure could not be ascribed to a recorded PC species. Control and treated groups showed 41 and 29 common species among them for porcine and human corneas, respectively. The unique PC species peaked at 12 minutes and at 30 minutes for human and porcine corneas followed by a decline consistent with an interplay of alkali penetration and hydrolyses at various time points. Conclusions. Alkali exposure dramatically changes the PC profile of cornea. Our data are consistent with penetration and hydrolysis as stochastic contributors to changes in PCs due to exposure to alkali for a finite duration and amount. PMID:22956606

The assessment of the stability of the corneal structure after LASIK correction of myopia by different optical zone diameters.

PubMed

Milivojević, Milorad; Petrović, Vladimir; Vukosavljević, Miroslav; Marjanović, Ivan; Resan, Mirko

2016-06-01

Enlargement of optical zone (OZ) diameter during laser in situ keratomileusis (LASIK) correction of myopia postoperatively improves the optical outcome, however, it also leads to the increased stroma tissue consumption--progressive corneal thinning. The aim of this investigation was to present the possibility of safe OZ enlargement without impairing the structural stability of the cornea, while obtaining an improved optical outcome with LASIK treatment of short-sightedness. Preoperative assessment of the cornea structure and prediction of the ablated stroma tissue consumption was conducted in 37 patients (74 eyes) treated for short-sightedness by means of the LASIK method. With the eyes that, according to their cornea structure, had the capacity for OZ diameter enlargement of 0.5 mm, LASIK treatment was performed within the wider OZ diameter of 7.0 mm compared to the standard 6.5 mm. The following two groups were formed, depending on the diameter of the utilized OZ: the group I (the eyes treated with the OZ 6.5 mm, n = 37) and the group II (the eyes treated with the OZ 7.0 mm, n = 37). No significant difference in the observed structural parameters of the cornea was detected between the groups of patients treated with different OZ diameters. The values of all the parameters were significantly bellow the threshold values for the development of postoperative ectasia. Diameter enlargement of the treated OZ, if there is a preoperative cornea capacity for such enlargement, will not impair the postoperative stability of the cornea structure, and will significantly improve the optical outcome.

Expression profiles of aquaporins in rat conjunctiva, cornea, lacrimal gland and Meibomian gland.

PubMed

Yu, Dongfang; Thelin, William R; Randell, Scott H; Boucher, Richard C

2012-10-01

The aim of the study was to elucidate aquaporin (AQP) family member mRNA expression and protein expression/localization in the rat lacrimal functional unit. The mRNA expression of all rat AQPs (AQP0-9, 11-12) in palpebral, fornical, and bulbar conjunctiva, cornea, lacrimal gland, and Meibomian gland was measured by Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) and real time RT-PCR. Antibodies against AQP1, 3, 4, 5, 9, and 11 were used in Western blotting and immunohistochemistry to determine protein expression and distribution. Our study demonstrated characteristic AQP expression profiles in rat ocular tissues. AQP1, 3, 4, 5, 8, 9, 11, and 12 mRNA were detected in conjunctiva. AQP0, 1, 2, 3, 4, 5, 6, 11, and 12 mRNA were expressed in cornea. AQP0, 1, 2, 3, 4, 5, 7, 8, and 11 mRNA were detected in lacrimal gland. AQP1, 3, 4, 5, 7, 8, 9, 11, and 12 mRNA were identified in Meibomian gland. By Western blot, AQP1, 3, 5, and 11 were detected in conjunctiva; AQP1, 3, 5, and 11 were identified in cornea; AQP1, 3, 4, 5, and 11 were detected in lacrimal gland; and AQP1, 3, 4, 5, 9, and 11 were present in Meibomian gland. Immunohistochemistry localized AQPs to distinct sites in the various tissues. This study rigorously analyzed AQPs expression and localization in rat conjunctiva, cornea, lacrimal gland, and Meibomian gland tissues. Our findings provide a comprehensive platform for further investigation into the physiological or pathophysiological relevance of AQPs in ocular surface. Copyright © 2012 Elsevier Ltd. All rights reserved.

Hypothermic preservation of corneas in a hyperkalaemic solution (CPTES): I. Short-term storage in the absence of colloid osmotic agents.

PubMed Central

Taylor, M J; Hunt, C J

1989-01-01

Preservation solutions for short-term storage of isolated donor corneas for use in penetrating keratoplasty have all been based on tissue culture medium, on the assumption that media designed to maintain the viability of cells at physiological temperatures will also provide suitable conditions for preservation at reduced temperatures. But for hypothermic preservation of some other tissues and organs, when ionic pumps are inhibited, it is unnecessary to support metabolism, and beneficial control of ion and water distribution between intra- and extracellular compartments is achieved by storage in appropriately formulated 'intracellular-type' solutions. We have therefore designed a solution that will restrict ionic imbalances and minimise endothelial cell swelling in corneas during exposure at reduced temperatures. This potassium-rich solution contains the biological pH buffer TES as an impermeant anion and is designated CPTES (corneal-potassium-TES). The structural and functional integrity of rabbit corneas stored at 0 degrees C in CPTES, without the addition of colloid osmotic agents, is compared with that of corneas stored in glutathione bicarbonate Ringers' solution (GBR), an 'extracellular-type' medium formulated for the maintenance of endothelial integrity during in-vitro perfusion at 34 degrees C. Corneas swelled significantly less during storage in CPTES than in GBR and could be stored for five days before reaching the same degree of hydration as corneas stored for only three days in GBR. Gross structural integrity and endothelial ultrastructure were maintained during storage for three and five days in CPTES. The rate of thinning of corneas stored in CPTES was significantly greater than in comparable groups of corneas stored in GBR. However, the efficient dehydration of corneas stored in CPTES was always preceded during perfusion by a brief period of additional swelling which was shown to be an osmotic response during the elution of the buffer compound TES that had permeated the stroma during storage. The omission of calcium or the addition of adenosine and glutathione to the CPTES preservation medium had no detectable effect on the integrity of the endothelium, but the omission of bicarbonate was beneficial, producing significantly higher rates of stromal thinning during normothermic perfusion. Additional benefits for extending storage by including colloid osmotic agents are described in a companion paper. Images PMID:2818988

Nondestructive quantification of analyte diffusion in cornea and sclera using optical coherence tomography.

PubMed

Ghosn, Mohamad G; Tuchin, Valery V; Larin, Kirill V

2007-06-01

Noninvasive functional imaging, monitoring, and quantification of analytes transport in epithelial ocular tissues are extremely important for therapy and diagnostics of many eye diseases. In this study the authors investigated the capability of optical coherence tomography (OCT) for noninvasive monitoring and quantification of diffusion of different analytes in sclera and cornea of rabbit eyes. A portable time-domain OCT system with wavelength of 1310 +/- 15 nm, output power of 3.5 mW, and resolution of 25 mum was used in this study. Diffusion of different analytes was monitored and quantified in rabbit cornea and sclera of whole eyeballs. Diffusion of water, metronidazole (0.5%), dexamethasone (0.2%), ciprofloxacin (0.3%), mannitol (20%), and glucose solution (20%) were examined, and their permeability coefficients were calculated by using OCT signal slope and depth-resolved amplitude methods. Permeability coefficients were calculated as a function of time and tissue depth. For instance, mannitol was found to have a permeability coefficient of (8.99 +/- 1.43) x 10(-6) cm/s in cornea and (6.18 +/- 1.08) x 10(-6) cm/s in sclera. The permeability coefficient of drugs with small concentrations (where water was the major solvent) was found to be in the range of that of water in the same tissue type, whereas permeability coefficients of higher concentrated solutions varied significantly. Results suggest that the OCT technique might be a powerful tool for noninvasive diffusion studies of different analytes in ocular tissues. However, additional methods of OCT signal acquisition and processing are required to study the diffusion of agents of small concentrations.

The grave necessity to make eye bank specular microscopy mandatory in all eye banks in the subcontinent to improve utilization of scarce donor corneas.

PubMed

Jadeja, Jagruti N; Patel, Bharati D; Shanbhag, Swapna S

2013-12-01

Donor tissue scarcity, Eye Bank Specular Microscopy as yet not made mandatory and tissue utilization often based on clinical judgment only. Prospectively analyze alteration in clinical grading of donor corneas and hence utilization, based on Eye Bank Specular Microscopy (EBSM) and to infer if EBSM should be mandatory in all eye banks. 200 consecutive otherwise 'suitable for surgery' donor eyes were graded clinically. On quantitative and qualitative analysis of endothelial cells by EBSM, final grading was adjusted. Impact on subsequent utilization for various surgeries was analyzed with regard to Age of Donor, Death to Enucleation Time, Death to Preservation Time and Lens Status of Donor Eye. 76 eyes (38%) (P < 0.05) had significant change in grading. 12/59 (20.30%) tissues from donors <=40 years showed Cell Density (CD) between 1801-2500. 19/76 (25%) tissues from donor >60 years showed CD >= 2500. From donor >=81 years, 2/13 (15.3%) eyes showed CD between 2501-3000 and 1 (7.6%) eye showed CD > 3000. Owing to better grading after EBSM, 13/14 (92.85%) tissues with DTET >6 hours and 5/5 (100%) tissues with DTPT > 16 hours were transplanted. Out of 45 (22.5%) pseudo-phakic tissues, 21 (46.67%) tissues were used for Therapeutic/Tectonic Penetrating Keratoplasty (PKP) while 24 (53.33%) tissues were used for Optical PKP. EBSM significantly alters final grading of tissues and its subsequent utilization. Acquiring huge importance in areas where adequate supply of corneas is lacking, EBSM becomes an indispensable tool for optimizing availability of qualified tissues for surgery. EBSM should be made a mandatory analysis.

Imaging of Keratoconic and normal human cornea with a Brillouin imaging system (Conference Presentation)

NASA Astrophysics Data System (ADS)

Besner, Sebastien; Shao, Peng; Scarcelli, Giuliano; Pineda, Roberto; Yun, Seok-Hyun (Andy)

2016-03-01

Keratoconus is a degenerative disorder of the eye characterized by human cornea thinning and morphological change to a more conical shape. Current diagnosis of this disease relies on topographic imaging of the cornea. Early and differential diagnosis is difficult. In keratoconus, mechanical properties are found to be compromised. A clinically available invasive technique capable of measuring the mechanical properties of the cornea is of significant importance for understanding the mechanism of keratoconus development and improve detection and intervention in keratoconus. The capability of Brillouin imaging to detect local longitudinal modulus in human cornea has been demonstrated previously. We report our non-contact, non-invasive, clinically viable Brillouin imaging system engineered to evaluate mechanical properties human cornea in vivo. The system takes advantage of a highly dispersive 2-stage virtually imaged phased array (VIPA) to detect weak Brillouin scattering signal from biological samples. With a 1.5-mW light beam from a 780-nm single-wavelength laser source, the system is able to detect Brillouin frequency shift of a single point in human cornea less than 0.3 second, at a 5μm/30μm lateral/axial resolution. Sensitivity of the system was quantified to be ~ 10 MHz. A-scans at different sample locations on a human cornea with a motorized human interface. We imaged both normal and keratoconic human corneas with this system. Whereas no significantly difference were observed outside keratocnic cones compared with normal cornea, a highly statistically significantly decrease was found in the cone regions.

Comparable change in stromal refractive index of cat and human corneas following blue-IRIS.

PubMed

Wozniak, Kaitlin T; Gearhart, Sara M; Savage, Daniel E; Ellis, Jonathan D; Knox, Wayne H; Huxlin, Krystel R

2017-05-01

Blue intratissue refractive index shaping (blue-IRIS) is a method with potential to correct ocular refraction noninvasively in humans. To date, blue-IRIS has only ever been applied to cat corneas and hydrogels. To test the comparability of refractive index change achievable in cat and human tissues, we used blue-IRIS to write identical phase gratings in ex vivo feline and human corneas. Femtosecond pulses (400 nm) were focused ? 300 ?? ? m below the epithelial surface of excised cat and human corneas and scanned to write phase gratings with lines ? 1 ?? ? m wide, spaced 5 ?? ? m apart, using a scan speed of 5 ?? mm / s . Additional cat corneas were used to test writing at 3 and 7 ?? mm / s in order to document speed dependence of the refractive index change magnitude. The first-order diffraction efficiency was immediately measured and used to calculate the refractive index change attained. Our data show that blue-IRIS induces comparable refractive index changes in feline and human corneas, an essential requirement for further developing its use as a clinical vision correction technique.

Comparable change in stromal refractive index of cat and human corneas following blue-IRIS

NASA Astrophysics Data System (ADS)

Wozniak, Kaitlin T.; Gearhart, Sara M.; Savage, Daniel E.; Ellis, Jonathan D.; Knox, Wayne H.; Huxlin, Krystel R.

2017-05-01

Blue intratissue refractive index shaping (blue-IRIS) is a method with potential to correct ocular refraction noninvasively in humans. To date, blue-IRIS has only ever been applied to cat corneas and hydrogels. To test the comparability of refractive index change achievable in cat and human tissues, we used blue-IRIS to write identical phase gratings in ex vivo feline and human corneas. Femtosecond pulses (400 nm) were focused ˜300 μm below the epithelial surface of excised cat and human corneas and scanned to write phase gratings with lines ˜1 μm wide, spaced 5 μm apart, using a scan speed of 5 mm/s. Additional cat corneas were used to test writing at 3 and 7 mm/s in order to document speed dependence of the refractive index change magnitude. The first-order diffraction efficiency was immediately measured and used to calculate the refractive index change attained. Our data show that blue-IRIS induces comparable refractive index changes in feline and human corneas, an essential requirement for further developing its use as a clinical vision correction technique.

In vivo 3D measurement of moxifloxacin and gatifloxacin distributions in the mouse cornea using multiphoton microscopy

NASA Astrophysics Data System (ADS)

Lee, Seunghun; Lee, Jun Ho; Park, Jin Hyoung; Yoon, Yeoreum; Chung, Wan Kyun; Tchah, Hungwon; Kim, Myoung Joon; Kim, Ki Hean

2016-05-01

Moxifloxacin and gatifloxacin are fourth-generation fluoroquinolone antibiotics used in the clinic to prevent or treat ocular infections. Their pharmacokinetics in the cornea is usually measured from extracted ocular fluids or tissues, and in vivo direct measurement is difficult. In this study multiphoton microscopy (MPM), which is a 3D optical microscopic technique based on multiphoton fluorescence, was applied to the measurement of moxifloxacin and gatifloxacin distribution in the cornea. Intrinsic multiphoton fluorescence properties of moxifloxacin and gatifloxacin were characterized, and their distributions in mouse cornea in vivo were measured by 3D MPM imaging. Both moxifloxacin and gatifloxacin had similar multiphoton spectra, while moxifloxacin had stronger fluorescence than gatifloxacin. MPM imaging of mouse cornea in vivo showed (1) moxifloxacin had good penetration through the superficial corneal epithelium, while gatifloxacin had relatively poor penetration, (2) both ophthalmic solutions had high intracellular distribution. In vivo MPM results were consistent with previous studies. This study demonstrates the feasibility of MPM as a method for in vivo direct measurement of moxifloxacin and gatifloxacin in the cornea.

Outcomes of shunt tube coverage with glycerol preserved cornea versus pericardium.

PubMed

Wigton, Eric; C Swanner, Jason; Joiner, Wade; Feldman, Alex; McGwin, Gerald; Huisingh, Carrie; Curcio, Christine A; Girkin, Christopher A

2014-01-01

Pericardium is a biomaterial widely used for covering the outflow tubes of glaucoma drainage devices. Recently, glycerol preserved cornea has been introduced as an alternative that offers durability and improved cosmesis because of its clarity. We retrospectively reviewed 262 patients in the University of Alabama Birmingham Glaucoma Service who underwent shunt procedures using either cornea tissue or pericardium to cover the tube. The primary outcome measure was the number of erosions of the covering material. Nine out of 101 (8.9%) patients in the pericardium covered group experienced an erosion compared with 3 out of 161 (1.9%) in the cornea covered group. A significant difference was reached with P=0.0125. Median follow-up was 440 days for the cornea group and 331 days for the pericardium group. The type of glaucoma (primary open-angle glaucoma vs. secondary glaucoma) was not associated with the risk of erosion (odds ratio, 0.501; 95% confidence interval, 0.204-1.234). The median time to exposure was 252 days in the pericardium group and 440 days in the cornea group (P=0.0017).

In vivo 3D measurement of moxifloxacin and gatifloxacin distributions in the mouse cornea using multiphoton microscopy

PubMed Central

Lee, Seunghun; Lee, Jun Ho; Park, Jin Hyoung; Yoon, Yeoreum; Chung, Wan Kyun; Tchah, Hungwon; Kim, Myoung Joon; Kim, Ki Hean

2016-01-01

Moxifloxacin and gatifloxacin are fourth-generation fluoroquinolone antibiotics used in the clinic to prevent or treat ocular infections. Their pharmacokinetics in the cornea is usually measured from extracted ocular fluids or tissues, and in vivo direct measurement is difficult. In this study multiphoton microscopy (MPM), which is a 3D optical microscopic technique based on multiphoton fluorescence, was applied to the measurement of moxifloxacin and gatifloxacin distribution in the cornea. Intrinsic multiphoton fluorescence properties of moxifloxacin and gatifloxacin were characterized, and their distributions in mouse cornea in vivo were measured by 3D MPM imaging. Both moxifloxacin and gatifloxacin had similar multiphoton spectra, while moxifloxacin had stronger fluorescence than gatifloxacin. MPM imaging of mouse cornea in vivo showed (1) moxifloxacin had good penetration through the superficial corneal epithelium, while gatifloxacin had relatively poor penetration, (2) both ophthalmic solutions had high intracellular distribution. In vivo MPM results were consistent with previous studies. This study demonstrates the feasibility of MPM as a method for in vivo direct measurement of moxifloxacin and gatifloxacin in the cornea. PMID:27138688

Organ Donation

MedlinePlus

Organ donation takes healthy organs and tissues from one person for transplantation into another. Experts say that the organs ... and bone marrow Cornea Most organ and tissue donations occur after the donor has died. But some ...

Optical coherence elastography assesses tissue modifications in laser reshaping of cornea and cartilages

NASA Astrophysics Data System (ADS)

Zaitsev, V. Y.; Matveyev, A. L.; Matveev, L. A.; Gelikonov, G. V.; Omelchenko, A. I.; Shabanov, D. V.; Sovetsky, A. A.; Baum, O. I.; Vitkin, A.; Sobol, E. N.

2018-02-01

Non-surgical thermo-mechanical reshaping of avascular collagenous tissues (cartilages and cornea) using moderate heating by IR-laser irradiation is an emerging technology that can find important applications in visioncorrection problems and preparation of cartilaginous implants in otolaryngology. To estimate both transient interframe strains and cumulative resultant strains produced by the laser irradiation of the tissue we use and improved version of strain mapping developed in our previous work related to compressional phase-sensitive optical coherence tomography. To reveal microstructural changes in the tissue regions where irradiation-produced strains do not disappear after temperature equilibration, we apply compressional optical coherence elastography in order to visualize the resultant variations in the tissue stiffness. The so-found regions of the stiffness reduction are attributed to formation of microscopic pores, existence of which agree with independent data obtained using methods of high-resolution microscopy.

UV cross-linking of donor corneas confers resistance to keratolysis.

PubMed

Arafat, Samer N; Robert, Marie-Claude; Shukla, Anita N; Dohlman, Claes H; Chodosh, James; Ciolino, Joseph B

2014-09-01

The aim of this study was to develop a modified ex vivo corneal cross-linking method that increases stromal resistance to enzymatic degradation for use as a carrier for the Boston keratoprosthesis. Ex vivo cross-linking of human corneas was performed using Barron artificial anterior chambers. The corneas were deepithelialized, pretreated with riboflavin solution (0.1% riboflavin/20% dextran), and irradiated with ultraviolet A (UV-A) light (λ = 370 nm, irradiance = 3 mW/cm) for various durations. The combined effect of UV-A and gamma (γ) irradiation was also assessed using the commercially available γ-irradiated corneal donors. The corneas were then trephined and incubated at 37°C with 0.3% collagenase A solution. The time to dissolution of each cornea was compared across treatments. Deepithelialized corneas (no UV light, no riboflavin) dissolved in 5.8 ± 0.6 hours. Cross-linked corneas demonstrated increased resistance to dissolution, with a time to dissolution of 17.8 ± 2.6 hours (P < 0.0001). The corneal tissues' resistance to collagenase increased with longer UV-A exposure, reaching a plateau at 30 minutes. Cross-linking both the anterior and posterior corneas did not provide added resistance when compared with cross-linking the anterior corneas only (P > 0.05). γ-irradiated corneas dissolved as readily as deepithelialized controls regardless of whether they were further cross-linked (5.6 ± 1.2 hours) or not (6.1 ± 0.6 hours) (P = 0.43). Collagen cross-linking of the deepithelialized anterior corneal surface for 30 minutes conferred optimal resistance to in vitro keratolysis by collagenase A.

Mechanisms of lamellar collagen formation in connective tissues.

PubMed

Ghazanfari, Samaneh; Khademhosseini, Ali; Smit, Theodoor H

2016-08-01

The objective of tissue engineering is to regenerate functional tissues. Engineering functional tissues requires an understanding of the mechanisms that guide the formation and evolution of structure in the extracellular matrix (ECM). In particular, the three-dimensional (3D) collagen fiber arrangement is important as it is the key structural determinant that provides mechanical integrity and biological function. In this review, we survey the current knowledge on collagen organization mechanisms that can be applied to create well-structured functional lamellar tissues and in particular intervertebral disc and cornea. Thus far, the mechanisms behind the formation of cross-aligned collagen fibers in the lamellar structures is not fully understood. We start with cell-induced collagen alignment and strain-stabilization behavior mechanisms which can explain a single anisotropically aligned collagen fiber layer. These mechanisms may explain why there is anisotropy in a single layer in the first place. However, they cannot explain why a consecutive collagen layer is laid down with an alternating alignment. Therefore, we explored another mechanism, called liquid crystal phasing. While dense concentrations of collagen show such behavior, there is little evidence that the conditions for liquid crystal phasing are actually met in vivo. Instead, lysyl aldehyde-derived collagen cross-links have been found essential for correct lamellar matrix deposition. Furthermore, we suggest that supra-cellular (tissue-level) shear stress may be instrumental in the alignment of collagen fibers. Understanding the potential mechanisms behind the lamellar collagen structure in connective tissues will lead to further improvement of the regeneration strategies of functional complex lamellar tissues. Copyright © 2016 Elsevier Ltd. All rights reserved.

New Technique of Exposed Glaucoma Drainage Tube Repair: Report of a Case.

PubMed

Berezina, Tamara L; Fechtner, Robert D; Cohen, Amir; Kim, Eliott E; Chu, David S

2015-01-01

We present the case of successful repair of an exposed glaucoma drainage tube by cornea graft fixation with tissue adhesive, and without subsequent coverage by adjacent conjunctiva or donor tissues. Patient with history of keratoglobus with thin cornea and sclera, and phthisical contralateral eye, underwent three unsuccessful corneal grafts followed by Boston type 1 keratoprosthesis in the right eye. Ahmed drainage device with sclera patch graft was implanted to control the intraocular pressure. Two years later the tube eroded through sclera graft and conjunctiva. Repair was performed by covering the tube with a corneal patch graft secured by tissue adhesive after the conjunctiva in this area was dissected away. The cornea graft was left uncovered due to fragility of adjacent conjunctiva. The healing of ocular and graft surfaces was complete prior to the 1 month follow-up. Conjunctival epithelium covered the corneal patch graft. At 12 months follow-up, the graft and the tube remained stable. Our report suggests that corneal patch graft fixation to the sclera by means of tissue adhesive, without closing the conjunctiva, can be considered as an effective alternative surgical approach for managing exposed glaucoma drainage tube, accompanied by adjacent conjunctiva tissue deficiency. How to cite this article: Berezina TL, Fechtner RD, Cohen A, Kim EE, Chu DS. New Technique of Exposed Glaucoma Drainage Tube Repair: Report of a Case. J Curr Glaucoma Pract 2015;9(2):62-64.

The Naïve Murine Cornea as a Model System to Identify Novel Endogenous Regulators of Lymphangiogenesis: TRAIL and rtPA.

PubMed

Regenfuß, Birgit; Dreisow, Marie-Luise; Hos, Deniz; Masli, Sharmila; Bock, Felix; Cursiefen, Claus

2015-06-01

In the murine cornea, which is an established model for analyzing pathologic lymphatic vessel growth, phenotypic heterogeneity of the endogenous lymphatic vessels in the limbus of the cornea was previously described. In this study, the cornea of BALB/c, C57BL/6, and FVB mice with different limbal lymphangiogenic phenotypes was analyzed to identify novel candidates potentially influencing lymphatic vessel growth. Pathway specific expression analysis of the cornea was performed to identify novel candidate genes. Corneal protein expression of the respective candidates was analyzed by fluorescent immunohistochemistry. The effect of the candidates on proliferation of human dermal lymphatic endothelial cells (HDLECs) was analyzed by BrdU proliferation ELISA. Thirteen genes were differentially regulated in corneas of mouse strains with more endogenous limbal lymphatic vessels (high-lymphangiogenic) (C57BL/6) compared to mouse strains with less endogenous limbal lymphatic vessels (low-lymphangiogenic) (BALB/c, FVB). Two candidates, Tumor necrosis factor (ligand) superfamily member 10 (Tnfsf10/Trail) and Plasminogen activator, tissue (Plat/tPA) were expressed in the cornea of BALB/c and C57BL/6 mice on the protein level. In vitro, Trail and recombinant tPA inhibited the proliferation of human dermal lymphatic endothelial cells. Molecular analysis of the naive cornea in mouse strains with different limbal lymphatic phenotypes is a valuable model to identify novel endogenous regulators of lymphangiogenesis.

Expression analysis of MDR1, BCRP and MRP3 transporter proteins in different in vitro and ex vivo cornea models for drug absorption studies.

PubMed

Verstraelen, Jessica; Reichl, Stephan

2013-01-30

Ocular drug absorption studies are required for the development of new drugs or drug delivery systems for eye treatment. Such preclinical investigations on transcorneal drug absorption are performed ex vivo with the excised corneas of experimental animals or in vitro using corneal cell culture models. The data currently available on the expression of ABC transporter proteins in corneal tissue is limited or contradictory. This study describes, for the first time, the comparison of the expression of ABC transporters, in particular, MDR1, BCRP and MRP3, between human cornea cell culture models and the most commonly used ex vivo models, namely, rabbit and porcine corneas, conducted in the same laboratory. The expression levels and functionality were determined by means of PCR, western blot, immunohistochemistry and bidirectional permeation studies using specific substrates and inhibitors. The results clearly indicate species-dependent expression of the studied efflux transporters. In the rabbit cornea, the expression and activity of MDR1 transporter was confirmed, whereas human cell culture models and porcine corneas did not show MDR1 expression. However, human cornea models possessed MRP3 and BCRP expression, whereas no functional expression was found in rabbit and porcine corneas. Therefore, the translation of transcorneal permeation data from animal experiments to humans should be performed with caution. Copyright © 2012 Elsevier B.V. All rights reserved.

BanTeC: a software tool for management of corneal transplantation.

PubMed

López-Alvarez, P; Caballero, F; Trias, J; Cortés, U; López-Navidad, A

2005-11-01

Until recently, all cornea information at our tissue bank was managed manually, no specific database or computer tool had been implemented to provide electronic versions of documents and medical reports. The main objective of the BanTeC project was therefore to create a computerized system to integrate and classify all the information and documents used in the center in order to facilitate management of retrieved, transplanted corneal tissues. We used the Windows platform to develop the project. Microsoft Access and Microsoft Jet Engine were used at the database level and Data Access Objects was the chosen data access technology. In short, the BanTeC software seeks to computerize the tissue bank. All the initial stages of the development have now been completed, from specification of needs, program design and implementation of the software components, to the total integration of the final result in the real production environment. BanTeC will allow the generation of statistical reports for analysis to improve our performance.

Femtosecond laser corneal surgery with in situ determination of the laser attenuation and ablation threshold by second harmonic generation

NASA Astrophysics Data System (ADS)

Plamann, Karsten; Nuzzo, Valeria; Albert, Olivier; Mourou, Gérard A.; Savoldelli, Michèle; Dagonet, Françoise; Donate, David; Legeais, Jean-Marc

2007-02-01

Femtosecond lasers start to be routinely used in refractive eye surgery. Current research focuses on their application to glaucoma and cataract surgery as well as cornea transplant procedures. To avoid unwanted tissue damage during the surgical intervention it is of utmost importance to maintain a working energy just above the ablation threshold and maintain the laser energy at this working point independently of the local and global tissue properties. To quantify the attenuation of the laser power density in the tissue by absorption, scattering and modification of the point spread function we monitor the second harmonic radiation generated in the collagen matrix of the cornea when exposed to ultrashort laser pulses. We use a CPA system with a regenerative amplifier delivering pulses at a wavelength of 1.06 μm, pulse durations of 400 fs and a maximum energy of 60 μJ. The repetition rate is adjustable from single shot up to 10 kHz. The experiments are performed on human corneas provided by the French Eye bank. To capture the SHG radiation we use a photomultiplier tube connected to a lockin amplifier tuned to the laser repetition rate. The measured data indicates an exponential decay of the laser beam intensity in the volume of the sample and allows for the quantification of the attenuation coefficient and its correlation with the optical properties of the cornea. Complementary analyses were performed on the samples by ultrastructural histology.

Photoablation of the cornea with a Q-switched Er:YAG laser

NASA Astrophysics Data System (ADS)

Lubatschowski, Holger; Hetzel, U.; Kermani, Omid; Ziolek, Carsten; Drommer, Wolfgang; Ertmer, Wolfgang

1997-12-01

In this study the ablation characteristics and the wound healing process of rabbit cornea irradiated with a Q- switched Er:YAG laser was evaluated. The laser, emitting at 2.94 micrometers wavelength, has a pulse width of 100 ns. The spot size on the corneal surface was 1 mm in diameter at a fluence of 750 mJ/cm2. The laser beam was applied by a `flying spot' mode, performing refractive ablations of -7 to -8 dpt. As a biological model, the corneas of 9 rabbits were irradiated. The post-treatment follow-up was as long as 39 days. The treated corneas were investigated by light and electron microscopy. The wound healing on rabbit cornea of the Q-switched Er:YAG laser radiation in corneal tissue processing resembles to what is known from ArF- excimer laser application. To shorten the pulse width by means of Q-switching is one major key to the successful application of the Er:YAG laser for PRK.

Implementation of Organ Culture storage of donor corneas: a 3 year study of its impact on the corneal transplant wait list at the Lions New South Wales Eye Bank.

PubMed

Devasahayam, Raj; Georges, Pierre; Hodge, Christopher; Treloggen, Jane; Cooper, Simon; Petsoglou, Con; Sutton, Gerard; Zhu, Meidong

2016-09-01

Organ Culture corneal storage offers an extended storage time and increased donor pool and tissue assessment opportunities. In September 2011, the Lions New South Wales Eye Bank (LNSWEB) moved from hypothermic storage to Organ Culture corneal storage. This study evaluates the impact of implementation of Organ Culture on donor eye retrieval and the corneal transplant waiting list over a 3 year period in NSW, Australia. Retrospective review of the LNSWEB data from September 2011 to August 2014. Tissue collection, waiting list and tissue utilization data were recorded. The data from September 2008 to August 2011 for Optisol-GS storage was used for comparison. The annual donor and cornea collection rate increased 35 % and 44 % respectively with Organ Culture compared to Optisol-GS storage. The utilization rate of corneal tissue increased from 73.4 % with hypothermic storage to 77.2 % with Organ Culture storage. The transplant wait list decreased by 77.3 % from September 2011 to August 2014 and correlated with the increased rate of corneal transplantation (r = -0.9381, p < 0.0001). No other factors impacting the wait list changed over this period. Corneas not used from either storage method were due to unacceptable endothelial cell density/viability. The contamination rate of corneas stored in Organ Culture medium was low at 1.74 %. The Organ Culture storage method increases the corneal donor pool available to Eye banks. The practical benefits of the extended storage time and increased donor assessment opportunities have directly led to an increase in corneal utilization rate and a significant decrease in recipient wait list time.

Absence of Vsx1 expression in the normal and damaged mouse cornea

PubMed Central

Chow, Robert L.

2011-01-01

Purpose To examine the expression of visual system homeobox 1 (Vsx1) in the mouse cornea and its potential role in the corneal wound response pathway. Methods Expression of Vsx1 was examined by quantitative reverse-transcription PCR (qRT–PCR) in corneal tissue from developing and adult mice and from mice that had undergone alkali-burn corneal wounding. Immunolabeling and Vsx1 knock-in reporter gene expression in wild type and Vsx1 null-mice were used to confirm the qRT–PCR data. Results Using qRT–PCR, Vsx1 expression was not detected in either the postnatal or adult mouse cornea or in corneas following wounding. This qRT–PCR data was supported by the absence of specific Vsx1 immunolabeling and Vsx1 knock-in reporter expression in untreated and wounded corneas. Conclusions In mice, Vsx1 mRNA, protein or reporter gene expression is not detected in the normal or damaged cornea. These results make it uncertain what role VSX1/Vsx1 plays in corneal biology. Future experiments examining the pathogenicity of VSX1 mutations associated with corneal dystrophy are required to rule out species differences and possible non-cell autonomous roles for VSX1 in the cornea. PMID:21437200

A quantitative method to evaluate the donor corneal tissue quality used in a comparative study between two hypothermic preservation media.

PubMed

Parekh, Mohit; Salvalaio, Gianni; Ferrari, Stefano; Amoureux, Marie-Claude; Albrecht, Cecile; Fortier, Denis; Ponzin, Diego

2014-12-01

To standardize a new evaluation technique for calculating the overall quality (OQ) of the donor cornea and validate it using a comparative study of corneas preserved in Optisol-GS and Cornea Cold®. Thirty pairs of donor corneas were selected for a 4 week in vitro comparative study using masked observers. Physiological parameters like thickness, transparency, viable endothelial cell density (VECD) and morphology were transformed to numerical range (0-4) to obtain the OQ. Microbiological examination was performed using Bactec instrument. Students t test showed statistically better results (p < 0.05) from week 3 for thickness, week 2 for transparency and week 1 for morphology and VECD; statistical significance (p < 0.05) was found for OQ from week 2 for the corneas preserved in Cornea Cold® compared to Optisol-GS. Epithelial quality was similar regardless of the medium. Microbiological examination showed absence of aerobic and anaerobic microorganisms in both media. OQ method is efficient, consistent and easy, now validated for comparative studies. Further refinement is necessary for its use at eye-banks, bio-banks and research or transplantation purposes. Cornea Cold® is a promising hypothermic corneal storage medium with preservation time ≤21 days. This permits higher flexibility, evaluation accuracy, longer duration for surgical preparation and ease of transportation.

Study of light scattering and transparency in human edematous corneas and application to corneal grafts

NASA Astrophysics Data System (ADS)

Marciano, Tal; Peyrot, Donald; Crotti, Caroline; Alahyane, Fatima; Kowalczuk, Laura; Plamann, Karsten

2011-07-01

The optical properties of the cornea have been a research subject of great interest for many years. Several early theories have been put forward to explain with more or less success the optical transparency of this tissue, but it was not until Maurice demonstrated in a very elegant way during the 50s that this optical transparency could be explained by the regular ultrastructure of the cornea. When becoming edematous, the cornea's ultrastructure is perturbed and the tissue becomes a strongly scattering medium. With the emergence of ophthalmologic surgery by ultrashort pulse lasers in recent years, a regain of interest in the subject of corneal transparency arose. However, relatively little and no recent data of transparency spectra measurements covering a large wavelength range is available in the literature. The purpose of this study is to provide quantitative values for light scattering and its relation to the degree of edema by measuring the spectrum of transmitted light through corneas presenting different degrees of edema. This paper focus on the comparison of laboratory measurements published earlier with a new simple method we propose We also for eye banks to quantitatively measure the degree of transparency of corneal grafts by measuring the modulation transfer function of a Siemens star viewed through a corneal graft. Indeed, there is no current method to determine the transparency of corneal graft but the subjectivity of the laboratory technician or the ophthalmic surgeon.

Intrastromal corneal reshaping using a high-intensity femtosecond laser: A novel method of vision correction

NASA Astrophysics Data System (ADS)

Han, Taehee

A new technology to perform a minimally invasive cornea reshaping procedure has been developed. This can eliminate the incidence of the flap-related complications of the conventional eye refractive procedures by multiphoton processes using a very high-intensity (I ≥ 1013 W/cm 2), but low energy (Ep ˜ 100-200 microJ) femtosecond laser pulses. Due to much lower energy than that of the nanosecond laser pulses for the thermal photoablation, the multiphoton processes cause almost no collateral damage by heat and shock wave generation. In this method, a series of femtosecond laser pulses is used to create very narrow (< 30 microm) and sufficiently long (≥ 2.5 mm) micro-channels in the cornea. The micro-channels are oriented almost perpendicular to the eye's optical axis. Once the micro-channel reaches a desired length, another series of femtosecond pulses with higher intensity is efficiently delivered through the micro-channel to the endpoint where a certain amount of the stromal tissue is disintegrated by the multiphoton processes. The disintegrated fragments are ejected out of the cornea via the same micro-channel, allowing the corneal surface to collapse, and changing its refractive power. This new corneal reshaping method obviates any process of damaging the corneal surface layer, while retaining the advantages of the conventional refractive procedures such as Laser in situ keratomileusis (LASIK) and Photorefractive keratectomy (PRK). In order to demonstrate the flapless cornea reshaping procedure, we have conducted ex-vivo experiments on fresh porcine eyes. The reshaped corneas were evaluated by using optical coherence tomography (OCT). The test results have shown that this flapless intrastromal procedure can reshape the cornea as intended with almost no surface damage. We have also performed a series of experiments to demonstrate the multiphoton processes in the corneal tissue by very high-intensity femtosecond laser pulses. Through the optical emission spectroscopy, we investigated the spectral lines of calcium atom and ions from the femtosecond laser-induced plasma from the porcine corneal tissue. The experimental results have shown the intensity-dependence of ablation rate, which qualitatively verifies the characteristics of the multiphoton processes.

Structure of corneal layers, collagen fibrils, and proteoglycans of tree shrew cornea.

PubMed

Almubrad, Turki; Akhtar, Saeed

2011-01-01

The stroma is the major part of the cornea, in which collagen fibrils and proteoglycans are distributed uniformly. We describe the ultrastructure of corneal layers, collagen fibrils (CF), and proteoglycans (PGs) in the tree shrew cornea. Tree shrew corneas (5, 6, and 10 week old animals) and normal human corneas (24, 25, and 54 years old) were fixed in 2.5% glutaraldehyde containing cuprolinic blue in a sodium acetate buffer. The tissue was processed for electron microscopy. The 'iTEM Olympus Soft Imaging Solutions GmbH' program was used to measure the corneal layers, collagen fibril diameters and proteoglycan areas. The tree shrew cornea consists of 5 layers: the epithelium, Bowman's layer, stroma, Descemet's membrane, and endothelium. The epithelium was composed of squamous cells, wing cells and basal cells. The Bowman's layer was 5.5±1.0 µm thick and very similar to a normal human Bowman's layer. The stroma was 258±7.00 µm thick and consisted of collagen fibril lamellae. The lamellae were interlaced with one another in the anterior stroma, but ran parallel to one another in the middle and posterior stroma. Collagen fibrils were decorated with proteoglycan filaments with an area size of 390 ±438 nm(2). The collagen fibril had a minimum diameter of 39±4.25 nm. The interfibrillar spacing was 52.91±6.07 nm. Within the collagen fibrils, very small electron-dense particles were present. The structure of the tree shrew cornea is very similar to that of the normal human cornea. As is the case with the human cornea, the tree shrew cornea had a Bowman's layer, lamellar interlacing in the anterior stroma and electron-dense particles within the collagen fibrils. The similarities of the tree shrew cornea with the human cornea suggest that it could be a good structural model to use when studying changes in collagen fibrils and proteoglycans in non-genetic corneal diseases, such as ectasia caused after LASIK (laser-assisted in situ keratomileusis).

Melatonin Entrains PER2::LUC Bioluminescence Circadian Rhythm in the Mouse Cornea

PubMed Central

Baba, Kenkichi; Davidson, Alec J.; Tosini, Gianluca

2015-01-01

Purpose Previous studies have reported the presence of a circadian rhythm in PERIOD2::LUCIFERASE (PER2::LUC) bioluminescence in mouse photoreceptors, retina, RPE, and cornea. Melatonin (MLT) modulates many physiological functions in the eye and it is believed to be one of the key circadian signals within the eye. The aim of the present study was to investigate the regulation of the PER2::LUC circadian rhythm in mouse cornea and to determine the role played by MLT. Methods Corneas were obtained from PER2::LUC mice and cultured to measure bioluminescence rhythmicity in isolated tissue using a Lumicycle or CCD camera. To determine the time-dependent resetting of the corneal circadian clocks in response to MLT or IIK7 (a melatonin type 2 receptor, MT2, agonist) was added to the cultured corneas at different times of the day. We also defined the location of the MT2 receptor within different corneal layers using immunohistochemistry. Results A long-lasting bioluminescence rhythm was recorded from cultured PER2::LUC cornea and PER2::LUC signal was localized to the corneal epithelium and endothelium. MLT administration in the early night delayed the cornea rhythm, whereas administration of MLT at late night to early morning advanced the cornea rhythm. Treatment with IIK7 mimicked the MLT phase-shifting effect. Consistent with these results, MT2 immunoreactivity was localized to the corneal epithelium and endothelium. Conclusions Our work demonstrates that MLT entrains the PER2::LUC bioluminescence rhythm in the cornea. Our data indicate that the cornea may represent a model to study the molecular mechanisms by which MLT affects the circadian clock. PMID:26207312

Effects of Microgravity on Quail Eye Development

NASA Technical Reports Server (NTRS)

Conrad, Gary W.

1996-01-01

During embryonic development, the most exposed tissue of the eye, the cornea, becomes differentially bulged outward because of constant intraocular pressure (IOP). The component cells of the cornea secrete a unique, paracrystalline extracellular matrix (the stroma) composed of orthogonal plies of collagen fibrils and proteoglycans. The cornea remains avascular, becomes transparent, and becomes more densely innervated than any other region on the surface of the body. Corneas from chicken embryos that flew on STS-47 contain many more cellular processes in the outermost region of the stroma (Bowman's Layer) than any corresponding region of control corneas. These processes appear to be cross-sections of cytoplasmic extensions of cells and are found in that region of Bowman's Layer immediately beneath the basal lamina of the corneal epithelium. Here, we propose to compare corneas of quail that flew in space on Mir-1 with those of ground controls to determine if the same unusual cellular processes are seen as in the space-flown chicken corneas. In the central regions of such space-flown corneas, the processes appear to be either portions of basal epithelial cells whose pseudopodial extensions have migrated down through their own basal lamina into the stroma, or corneal nerves that have innervated the corneal stroma in an unusual manner. Eyeballs of embryos fixed on Mir-1, control embryos fixed at KSC and clinostated embryos fixed at KSU, will provide corneas for this study. Electron microscopy will be used to assess the distribution of the cellular processes in Bowman's Layer in the central region of each cornea. Attempts also will be made to determine the relative glycosaminoglycan distributions in the corneal stromas by indirect immunofluorscence and to record whole-mount staining patterns of the corneal nerves.

Optical coherence tomography in estimating molecular diffusion of drugs and analytes in ocular tissues

NASA Astrophysics Data System (ADS)

Ghosn, Mohamad G.; Tuchin, Valery V.; Larin, Kirill V.

2009-02-01

Aside from other ocular drug delivery methods, topical application and follow up drug diffusion through the cornea and sclera of the eye remain the favored method, as they impose the least pain and discomfort to the patient. However, this delivery route suffers from the low permeability of epithelial tissues and drug washout, thus reducing the effectiveness of the drug and ability to reach its target in effective concentrations. In order to better understand the behavioral characteristics of diffusion in ocular tissue, a method for noninvasive imaging of drug diffusion is needed. Due to its high resolution and depth-resolved imaging capabilities, optical coherence tomography (OCT) has been utilized in quantifying the molecular transport of different drugs and analytes in vitro in the sclera and the cornea. Diffusion of Metronidazole (0.5%), Dexamethasone (0.2%), Ciprofloxacin (0.3%), Mannitol (20%), and glucose solution (20%) in rabbit sclera and cornea were examined. Their permeability coefficients were calculated by using OCT signal slope and depth-resolved amplitude methods as function of time and tissue depth. For instance, mannitol was found to have a permeability coefficient of (8.99 +/- 1.43) × 10-6 cm/s in cornea (n=4) and (6.18 +/- 1.08) × 10-6 cm/s in sclera (n=5). We also demonstrate the capability of OCT technique for depth-resolved monitoring and quantifying of glucose diffusion in different layers of the sclera. We found that the glucose diffusion rate is not uniform throughout the tissue and is increased from approximately (2.39 +/- 0.73) × 10-6 cm/s at the epithelial side to (8.63 +/- 0.27) × 10-6 cm/s close to the endothelial side of the sclera. In addition, discrepancy in the permeability rates of glucose solutions with different concentrations was observed. Such diffusion studies could enhance our knowledge and potentially pave the way for advancements of therapeutic and diagnostic techniques in the treatment of ocular diseases.

Effect of dehydration in the UV transmittance of "in vitro" corneas

NASA Astrophysics Data System (ADS)

Lincoln, Victor A. C.; Ventura, Liliane; Faria e Sousa, Sidney J.; Mello, Marcio M.

2012-03-01

In ophthalmology the research using "in vitro" corneas are an excellent model for studies of new ophthalmologic procedures, enabling the analysis of effectiveness, performance and even safety parameters of the procedure. In this work we studied four "in vitro" human corneas preserved in OPTISOL-GS, with initial average pachymetry of 542 microns and a post-mortem average of 6 days. The corneas were preserved in OPTISOL-GS and were washed with saline solution to remove the excess the preservative medium. The corneas were placed in a device aligned with an ultraviolet source of 3mw/cm2 and an optical fiber positioned after the device near the endothelium of the cornea. The UV transmittance spectra in the region of 360-370nm were captured by the emission of UV source for 3 seconds. These spectra were captured every 5 minutes in a total of 60 minutes, resulting in 13 spectra per cornea. The measured average initial UV transmittance was 73% and after 50 minutes of dehydration there was no significant difference in the corneal teansmittance properties. However, for the last 10 minutes we have observed a decrease in the UV transmittance of 4%, probably indicated by corneal dehydration and swelling (wrinkling of the cornea tissue. The final average pachymetry was 421 microns and the UV transmittance after the 60 minutes was 69%. Therefore we can suppose that the UV transmittance of corneas "in vitro" is invariant over a period of up to 60 minutes, even with the thickness decrease, since the material that absorbs in the UV region remains intact and only water loss occurs.

An improved biofunction of titanium for keratoprosthesis by hydroxyapatite-coating.

PubMed

Dong, Ying; Yang, Jingxin; Wang, Liqiang; Ma, Xiao; Huang, Yifei; Qiu, Zhiye; Cui, Fuzhai

2014-03-01

Titanium framework keratoprosthesis has been commonly used in the severe corneal blindness, but the tissue melting occurred frequently around titanium. Since hydroxyapatite has been approved to possess a good tissue integration characteristic, nanostructured hydroxyapatite was coated on the surface of titanium through the aerosol deposition method. In this study, nanostructured hydroxyapatite coating was characterized by X-ray diffraction, scanning electron microscopy, atomic force microscopy, and auger electronic spectrometer. Biological evaluations were performed with rabbit cornea fibroblast in vitro and an animal model in vivo. The outcomes showed the coating had a grain-like surface topography and a good atomic mixed area with substrate. The rabbit cornea fibroblasts appeared a good adhesion on the surface of nanostructured hydroxyapatite in vitro. In the animal model, nanostructured hydroxyapatite-titanium implants were stably retained in the rabbit cornea, and by contrast, the corneal stroma became thinner anterior to the implants in the control. Therefore, our findings proved that nanostructured hydroxyapatite-titanium could not only provide an improved bond for substrate but also enhance the tissue integration with implants in host. As a promising material, nanostructured hydroxyapatite-titanium-based keratoprosthesis prepared by the aerosol deposition method could be utilized for the corneal blindness treatment.

Lens regeneration from the cornea requires suppression of Wnt/β-catenin signaling.

PubMed

Hamilton, Paul W; Sun, Yu; Henry, Jonathan J

2016-04-01

The frog, Xenopus laevis, possesses a high capacity to regenerate various larval tissues, including the lens, which is capable of complete regeneration from the cornea epithelium. However, the molecular signaling mechanisms of cornea-lens regeneration are not fully understood. Previous work has implicated the involvement of the Wnt signaling pathway, but molecular studies have been very limited. Iris-derived lens regeneration in the newt (Wolffian lens regeneration) has shown a necessity for active Wnt signaling in order to regenerate a new lens. Here we provide evidence that the Wnt signaling pathway plays a different role in the context of cornea-lens regeneration in Xenopus. We examined the expression of frizzled receptors and wnt ligands in the frog cornea epithelium. Numerous frizzled receptors (fzd1, fzd2, fzd3, fzd4, fzd6, fzd7, fzd8, and fzd10) and wnt ligands (wnt2b.a, wnt3a, wnt4, wnt5a, wnt5b, wnt6, wnt7b, wnt10a, wnt11, and wnt11b) are expressed in the cornea epithelium, demonstrating that this tissue is transcribing many of the ligands and receptors of the Wnt signaling pathway. When compared to flank epithelium, which is lens regeneration incompetent, only wnt11 and wnt11b are different (present only in the cornea epithelium), identifying them as potential regulators of cornea-lens regeneration. To detect changes in canonical Wnt/β-catenin signaling occurring within the cornea epithelium, axin2 expression was measured over the course of regeneration. axin2 is a well-established reporter of active Wnt/β-catenin signaling, and its expression shows a significant decrease at 24 h post-lentectomy. This decrease recovers to normal endogenous levels by 48 h. To test whether this signaling decrease was necessary for lens regeneration to occur, regenerating eyes were treated with either 6-bromoindirubin-3'-oxime (BIO) or 1-azakenpaullone - both activators of Wnt signaling - resulting in a significant reduction in the percentage of cases with successful regeneration. In contrast, inhibition of Wnt signaling using either the small molecule IWR-1, treatment with recombinant human Dickkopf-1 (rhDKK1) protein, or transgenic expression of Xenopus DKK1, did not significantly affect the percentage of successful regeneration. Together, these results suggest a model where Wnt/β-catenin signaling is active in the cornea epithelium and needs to be suppressed during early lens regeneration in order for these cornea cells to give rise to a new lentoid. While this finding differs from what has been described in the newt, it closely resembles the role of Wnt signaling during the initial formation of the lens placode from the surface ectoderm during early embryogenesis. Copyright © 2016 Elsevier Ltd. All rights reserved.

Portable light transmission measuring system for preserved corneas.

PubMed

Ventura, Liliane; Jesus, Gabriel Torres de; Oliveira, Gunter Camilo Dablas de; Sousa, Sidney J F

2005-12-22

The authors have developed a small portable device for the objective measurement of the transparency of corneas stored in preservative medium, for use by eye banks in evaluation prior to transplantation. The optical system consists of a white light, lenses, and pinholes that collimate the white light beams and illuminate the cornea in its preservative medium, and an optical filter (400-700 nm) that selects the range of the wavelength of interest. A sensor detects the light that passes through the cornea, and the average corneal transparency is displayed. In order to obtain only the tissue transparency, an electronic circuit was built to detect a baseline input of the preservative medium prior to the measurement of corneal transparency. The operation of the system involves three steps: adjusting the "0 %" transmittance of the instrument, determining the "100 %" transmittance of the system, and finally measuring the transparency of the preserved cornea inside the storage medium. Fifty selected corneas were evaluated. Each cornea was submitted to three evaluation methods: subjective classification of transparency through a slit lamp, quantification of the transmittance of light using a corneal spectrophotometer previously developed, and measurement of transparency with the portable device. By comparing the three methods and using the expertise of eye bank trained personnel, a table for quantifying corneal transparency with the new device has been developed. The correlation factor between the corneal spectrophotometer and the new device is 0,99813, leading to a system that is able to standardize transparency measurements of preserved corneas, which is currently done subjectively.

Effects of the holmium laser on the human cornea: a preliminary study

NASA Astrophysics Data System (ADS)

Mueller, Linda J.; Tassignon, Marie J.; Trau, Rene; Pels, Liesbeth; Vrensen, Gijs F.

1996-12-01

Treatment of peripheral post-mortem human corneas with the Holmium laser in a ring pattern resulted in opaque spots. One pair of treated eyes was immediately processed for light and electron microscopy and three other treated eyes were preserved for 4 days in medium in order to compare direct and short-term effects of the Holmium laser. Cross as well as frontal light microscopical sections of all eyes revealed interconnecting bands between the spots. At the ultrastructural level the anterior corneal tissue within these spots was characterized by coagulation of cells and collagen and shoed either a dramatic distorting effect on the epithelium in the eyes processed immediately or a single layer of flattened multi-nucleolated epithelial cells having more than one nucleolus per nucleus in the eyes stored in medium. Furthermore, the spots showed disturbed Bowman's layer, destroyed keratocytes and collagen fibrils which were either coagulated or organized chaotically. The interconnecting bands contained alternating normal and coagulated collagen fibers. The rest of the cornea outside the spots had a normal appearance. In corneas stored in medium, both keratocytes and epithelial cells over the entire cornea exhibited accumulations of cytoplasmic fibrils and glycogen particles. These phenomena were not observed in non-preserved corneas, suggesting that the differences are due to preservation and not due to the laser treatment. It is concluded that morphological changes occur mainly in the treated peripheral cornea whereas the central untreated cornea remains unaffected,indicating that the Holmium laser is a reliable instrument to treat hypermetropic patients.

Combined use of a femtosecond laser and a microkeratome in obtaining thin grafts for Descemet stripping automated endothelial keratoplasty: an eye bank study.

PubMed

Murta, Joaquim N; Rosa, Andreia M; Quadrado, Maria Joao C; Russo, Ana D; Brito, Sergio S; Silva, Maria Fátima L

2013-01-01

To evaluate the use of a femtosecond laser combined with a microkeratome in the preparation of posterior corneal disks for Descemet stripping automated endothelial keratoplasty (DSAEK). This experimental study involved ultrathin DSAEK tissue preparation of 22 donor corneas unsuitable for transplantation. The first cut was performed with an Intralase® FS60 laser and the second cut with a Moria CBm 300-µm microkeratome. The thickness of the first cut was modified for each cornea to obtain a final graft thickness of less than 110 µm. Precut and postcut central pachymetry were performed with an ultrasonic pachymeter. Central endothelial cell density (ECD) was calculated before and 24 hours after tissue preparation. Final graft thickness was 105.0 ± 26.1 (SD) µm (range 65-117). The mean microkeratome head cut thickness was 324.5 ± 10.9 µm (range 310-345). Precut and postcut ECDs averaged 2250 ± 222 and 2093 ± 286 cells/mm2, respectively, representing 6.9% of cell loss. No corneas were perforated. Femtosecond FS60 lasers and Moria CBm 300-µm microkeratomes can be used sequentially to prepare consistently thin DSAEK grafts with no irregular cuts or cornea perforations.

Gene Therapy in the Cornea: 2005-present

PubMed Central

Mohan, Rajiv R.; Tovey, Jonathan C.K.; Sharma, Ajay; Tandon, Ashish

2011-01-01

Successful restoration of vision in human patients with gene therapy affirmed its promise to cure ocular diseases and disorders. The efficacy of gene therapy is contingent upon vector and mode of therapeutic DNA introduction into targeted cells/tissues. The cornea is an ideal tissue for gene therapy due to its ease of access and relative immune-privilege. Considerable progress has been made in the field of corneal gene therapy in last 5 years. Several new gene transfer vectors, techniques and approaches have evolved. Although corneal gene therapy is still in its early stages of development, the potential of gene-based interventions to treat corneal abnormalities have begun to surface. Identification of next generation viral and nanoparticle vectors, characterization of delivered gene levels, localization, and duration in the cornea, and significant success in controlling corneal disorders, particularly fibrosis and angiogenesis, in experimental animal disease models, with no major side effects have propelled gene therapy a step closer towards establishing gene-based therapies for corneal blindness. Recently, researchers have assessed the delivery of therapeutic genes for corneal diseases and disorders due to trauma, infections, chemical, mechanical, and surgical injury, and/or abnormal wound healing. This review provides an update on the developments in gene therapy for corneal diseases and discusses the barriers that hinder its utilization for delivering genes in the cornea. PMID:21967960

Determining the Depth of Injury in Bioengineered Tissue Models of Cornea and Conjunctiva for the Prediction of All Three Ocular GHS Categories

PubMed Central

Zorn-Kruppa, Michaela; Houdek, Pia; Wladykowski, Ewa; Engelke, Maria; Bartok, Melinda; Mewes, Karsten R.; Moll, Ingrid; Brandner, Johanna M.

2014-01-01

The depth of injury (DOI) is a mechanistic correlate to the ocular irritation response. Attempts to quantitatively determine the DOI in alternative tests have been limited to ex vivo animal eyes by fluorescent staining for biomarkers of cell death and viability in histological cross sections. It was the purpose of this study to assess whether DOI could also be measured by means of cell viability detected by the MTT assay using 3-dimensional (3D) reconstructed models of cornea and conjunctiva. The formazan-free area of metabolically inactive cells in the tissue after topical substance application is used as the visible correlate of the DOI. Areas of metabolically active or inactive cells are quantitatively analyzed on cryosection images with ImageJ software analysis tools. By incorporating the total tissue thickness, the relative MTT-DOI (rMTT-DOI) was calculated. Using the rMTT-DOI and human reconstructed cornea equivalents, we developed a prediction model based on suitable viability cut-off values. We tested 25 chemicals that cover the whole range of eye irritation potential based on the globally harmonized system of classification and labelling of chemicals (GHS). Principally, the MTT-DOI test method allows distinguishing between the cytotoxic effects of the different chemicals in accordance with all 3 GHS categories for eye irritation. Although the prediction model is slightly over-predictive with respect to non-irritants, it promises to be highly valuable to discriminate between severe irritants (Cat. 1), and mild to moderate irritants (Cat. 2). We also tested 3D conjunctiva models with the aim to specifically address conjunctiva-damaging substances. Using the MTT-DOI method in this model delivers comparable results as the cornea model, but does not add additional information. However, the MTT-DOI method using reconstructed cornea models already provided good predictability that was superior to the already existing established in vitro/ex vivo methods. PMID:25494045

Evaluation of active and passive transport processes in corneas extracted from preserved rabbit eyes.

PubMed

Majumdar, Soumyajit; Hingorani, Tushar; Srirangam, Ramesh

2010-04-01

In vitro transcorneal permeability studies are an important screening tool in drug development. The objective of this research is to examine the feasibility of using corneas isolated from preserved rabbit eyes as a model for permeability evaluation. Eyes from male New Zealand White rabbits were used immediately or were stored overnight in phosphate-buffered saline (PBS) or Hanks balanced salt solution (HBSS) over wet ice. Integrity of isolated corneas was evaluated by measuring the TEER and by determining the permeability of paracellular and transcellular markers. Active transport was assessed by measuring transcorneal permeability of selected amino acids. Esterase activity was estimated using p-nitrophenyl assay. In all cases, corneas from freshly enucleated eyes were compared to those isolated from the day-old preserved eyes. Transcellular and paracellular passive diffusion was not affected by the storage medium and observed to be similar in the fresh and preserved eye models. However, amino acid transporters demonstrated lower functional activity in corneas excised from eyes preserved in PBS. Moreover, preserved eyes displayed almost 1.5-fold lower esterase activity in the corneal tissue. Thus, corneas isolated from day-old eyes, preserved in HBSS, closely mimics freshly excised rabbit corneas in terms of both active and passive transport characteristics but possesses slightly reduced enzymatic activity. 2009 Wiley-Liss, Inc. and the American Pharmacists Association

Terahertz time-lapse imaging of hydration in physiological tissues

NASA Astrophysics Data System (ADS)

Bennett, David B.; Taylor, Zachary D.; Bajwa, Neha; Tewari, Priyamvada; Maccabi, Ashkan; Sung, Shijun; Singh, Rahul S.; Culjat, Martin O.; Grundfest, Warren S.; Brown, Elliott R.

2011-02-01

This study describes terahertz (THz) imaging of hydration changes in physiological tissues with high water concentration sensitivity. A fast-scanning, pulsed THz imaging system (centered at 525 GHz; 125 GHz bandwidth) was utilized to acquire a 35 mm x 35 mm field-of-view with 0.5 mm x 0.5 mm pixels in less than two minutes. THz time-lapsed images were taken on three sample systems: (1) a simple binary system of water evaporating from a polypropylene towel, (2) the accumulation of fluid at the site of a sulfuric acid burn on ex vivo porcine skin, and (3) the evaporative dehydration of an ex vivo porcine cornea. The diffusion-regulating behavior of corneal tissue is elucidated, and the correlation of THz reflectivity with tissue hydration is measured using THz spectroscopy on four ex vivo corneas. We conclude that THz imaging can discern small differences in the distribution of water in physiological tissues and is a good candidate for burn and corneal imaging.

Occurrence of viral DNA in paired samples of corneal rim and cornea preservation fluid.

PubMed

Broniek, G; Langwińska-Wośko, E; Sybilska, M; Szaflik, J P; Przybylski, M; Wróblewska, M

2017-04-01

Corneal transplants have one of the highest success rates among all transplantological procedures. Corneas intended for transplantation are stored in a preservation fluid, which is then tested for bacterial and fungal infections. Among all analyses of infectious complications following corneal transplants, infections caused by bacteria or fungi are the most prominent. Surprisingly, however, apart from a few publications, there is a lack of data regarding the occurrence of viruses in donor corneas and the risk of transmitting these to their recipients. The intention of this research was therefore to determine the frequency with which human herpesvirus 1 (HHV-1), human herpesvirus 2 (HHV-2), and human adenovirus (HAdV) occur in transplanted corneal tissue, as well as in samples of preservation fluid. The study comprised 57 paired samples, with each pair consisting of a fragment of the corneal tissue remaining after its trepanation for transplantation surgery and a sample of corneal preservation fluid. Sample pairs were all tested for the presence of the DNA of three viruses (HHV-1, HHV-2, and HAdV) using real time PCR technique. Viral DNA was found in three of the tested corneas-HHV-1 DNA in one paired sample (1.8%) and adenovirus DNA in two single samples (3.5%). We postulate that virological testing of corneas for transplantation should be considered, particularly in the case of donors with increased risk factors for herpesvirus and adenovirus reactivation. J. Med. Virol. 89:732-736, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Ocular HSV-1: Is the Cornea a Reservoir for Viral Latency or a Fast Pit Stop?

PubMed Central

Kennedy, David P.; Clement, Christian; Arceneaux, Richard L.; Bhattacharjee, Partha S.; Huq, Tashfin S.; Hill, James M.

2010-01-01

Purpose To present a review supporting and refuting evidence from mouse, rabbit, non-human primate, and human studies of herpes simplex virus type 1 (HSV-1) concerning corneal latency. Methods More than 50 research papers on HSV-1 published in peer-reviewed journals were examined. Results Infectious HSV-1 has been found in mouse denervated tissues and in tissues with negative cultures from the corresponding ganglion. However, the different mouse strains have shown varied responses to different strains of HSV, making it difficult to relate such findings to humans. Rabbit studies provide excellent evidence for HSV-1 corneal latency including data on HSV-1 migration from the cornea into the corneoscleral rim and on the distribution of HSV-1 DNA in the cornea. However, the available methods for the detection of infectious HSV-1 may not be sensitive enough to detect low-level infection. Infectious HSV-1 has been successfully isolated from the tears of non-human primates in the absence of detectable corneal lesions. The recurrence of corneal ulcers in non-human primates before the appearance of infectious HSV-1 in tears suggests that the origin of the HSV-1 is the cornea, rather than the TG. Human studies presented evidence of both ganglion and corneal latency. Conclusion Understanding HSV-1 disease progression and the possibility of corneal latency could lead to more effective treatments for herpetic keratitis. However, it is unlikely that operational latency in the cornea will be definitively proven unless a new method with higher sensitivity for the detection of infectious virus is developed. PMID:21304287

The corneal transplant score: a simple corneal graft candidate calculator.

PubMed

Rosenfeld, Eldar; Varssano, David

2013-07-01

Shortage of corneas for transplantation has created long waiting lists in most countries. Transplant calculators are available for many organs. The purpose of this study is to describe a simple automatic scoring system for keratoplasty recipient candidates, based on several parameters that we consider most relevant for tissue allocation, and to compare the system's accuracy in predicting decisions made by a cornea specialist. Twenty pairs of candidate data were randomly created on an electronic spreadsheet. A single priority score was computed from the data of each candidate. A cornea surgeon and the automated system then decided independently which candidate in each pair should have surgery if only a single cornea was available. The scoring system can calculate values between 0 (lowest priority) and 18 (highest priority) for each candidate. Average score value in our randomly created cohort was 6.35 ± 2.38 (mean ± SD), range 1.28 to 10.76. Average score difference between the candidates in each pair was 3.12 ± 2.10, range 0.08 to 8.45. The manual scoring process, although theoretical, was mentally and emotionally demanding for the surgeon. Agreement was achieved between the human decision and the calculated value in 19 of 20 pairs. Disagreement was reached in the pair with the lowest score difference (0.08). With worldwide donor cornea shortage, waiting for transplantation can be long. Manual sorting of priority for transplantation in a long waiting list is difficult, time-consuming and prone to error. The suggested system may help achieve a justified distribution of available tissue.

Two-photon spectral fluorescence lifetime and second-harmonic generation imaging of the porcine cornea with a 12-femtosecond laser microscope

NASA Astrophysics Data System (ADS)

Batista, Ana; Breunig, Hans Georg; Uchugonova, Aisada; Morgado, António Miguel; König, Karsten

2016-03-01

Five dimensional microscopy with a 12-fs laser scanning microscope based on spectrally resolved two-photon autofluorescence lifetime and second-harmonic generation (SHG) imaging was used to characterize all layers of the porcine cornea. This setup allowed the simultaneous excitation of both metabolic cofactors, NAD(P)H and flavins, and their discrimination based on their spectral emission properties and fluorescence decay characteristics. Furthermore, the architecture of the stromal collagen fibrils was assessed by SHG imaging in both forward and backward directions. Information on the metabolic state and the tissue architecture of the porcine cornea were obtained with subcellular resolution, and high temporal and spectral resolutions.

Two-photon spectral fluorescence lifetime and second-harmonic generation imaging of the porcine cornea with a 12-femtosecond laser microscope.

PubMed

Batista, Ana; Breunig, Hans Georg; Uchugonova, Aisada; Morgado, António Miguel; König, Karsten

2016-03-01

Five dimensional microscopy with a 12-fs laser scanning microscope based on spectrally resolved two-photon autofluorescence lifetime and second-harmonic generation (SHG) imaging was used to characterize all layers of the porcine cornea. This setup allowed the simultaneous excitation of both metabolic cofactors, NAD(P)H and flavins, and their discrimination based on their spectral emission properties and fluorescence decay characteristics. Furthermore, the architecture of the stromal collagen fibrils was assessed by SHG imaging in both forward and backward directions. Information on the metabolic state and the tissue architecture of the porcine cornea were obtained with subcellular resolution, and high temporal and spectral resolutions.

Corneal avascularity is due to soluble VEGF receptor-1

PubMed Central

Ambati, Balamurali K.; Nozaki, Miho; Singh, Nirbhai; Takeda, Atsunobu; Jani, Pooja D.; Suthar, Tushar; Albuquerque, Romulo J. C.; Richter, Elizabeth; Sakurai, Eiji; Newcomb, Michael T.; Kleinman, Mark E.; Caldwell, Ruth B.; Lin, Qing; Ogura, Yuichiro; Orecchia, Angela; Samuelson, Don A.; Agnew, Dalen W.; Leger, Judy St.; Green, W. Richard; Mahasreshti, Parameshwar J.; Curiel, David T.; Kwan, Donna; Marsh, Helene; Ikeda, Sakae; Leiper, Lucy J.; Collinson, J. Martin; Bogdanovich, Sasha; Khurana, Tejvir S.; Shibuya, Masabumi; Baldwin, Megan E.; Ferrara, Napoleone; Gerber, Hans-Peter; Falco, Sandro De; Witta, Jassir; Baffi, Judit Z.; Raisler, Brian J.; Ambati, Jayakrishna

2009-01-01

Corneal avascularity—the absence of blood vessels in the cornea—is required for optical clarity and optimal vision, and has led to the cornea being widely used for validating pro- and anti-angiogenic therapeutic strategies for many disorders1-4. But the molecular underpinnings of the avascular phenotype have until now remained obscure5-10 and are all the more remarkable given the presence in the cornea of vascular endothelial growth factor (VEGF)-A, a potent stimulator of angiogenesis, and the proximity of the cornea to vascularized tissues. Here we show that the cornea expresses soluble VEGF receptor-1 (sVEGFR-1; also known as sflt-1) and that suppression of this endogenous VEGF-A trap11 by neutralizing antibodies, RNA interference or Cre-lox-mediated gene disruption abolishes corneal avascularity in mice. The spontaneously vascularized corneas of corn1 and Pax6+/− mice12,13 and Pax6+/− patients with aniridia14 are deficient in sflt-1, and recombinant sflt-1 administration restores corneal avascularity in corn1 and Pax6+/− mice. Manatees, the only known creatures uniformly to have vascularized corneas15, do not express sflt-1, whereas the avascular corneas of dugongs, also members of the order Sirenia, elephants, the closest extant terrestrial phylogenetic relatives of manatees, and other marine mammals (dolphins and whales) contain sflt-1, indicating that it has a crucial, evolutionarily conserved role. The recognition that sflt-1 is essential for preserving the avascular ambit of the cornea can rationally guide its use as a platform for angiogenic modulators, supports its use in treating neovascular diseases, and might provide insight into the immunological privilege of the cornea. PMID:17051153

Engineering a collagen matrix that replicates the biological properties of native extracellular matrix.

PubMed

Nam, Kwangwoo; Sakai, Yuuki; Funamoto, Seiichi; Kimura, Tsuyoshi; Kishida, Akio

2011-01-01

In this study, we aimed to replicate the function of native tissues that can be used in tissue engineering and regenerative medicine. The key to such replication is the preparation of an artificial collagen matrix that possesses a structure resembling that of the extracellular matrix. We, therefore, prepared a collagen matrix by fibrillogenesis in a NaCl/Na(2)HPO(4) aqueous solution using a dialysis cassette and investigated its biological behavior in vitro and in vivo. The in vitro cell adhesion and proliferation did not show any significant differences. The degradation rate in the living body could be controlled according to the preparation condition, where the collagen matrix with high water content (F-collagen matrix, >98%) showed fast degradation and collagen matrix with lower water content (T-collagen matrix, >80%) showed no degradation for 8 weeks. The degradation did not affect the inflammatory response at all and relatively faster wound healing response was observed. Comparing this result with that of collagen gel and decellularized cornea, it can be concluded that the structural factor is very important and no cell abnormal behavior would be observed for quaternary structured collagen matrix.

Quantification of phase retardation in corneal tissues using a femtosecond laser

NASA Astrophysics Data System (ADS)

Calhoun, William R.; Beylin, Alexander; Weiblinger, Richard; Ilev, Ilko

2013-03-01

The use of femtosecond lasers (FSL) in ophthalmic procedures, such as LASIK, lens replacement (cataract surgery), as well as several other treatments, is growing rapidly. The treatment effect is based on photo ablation of ocular tissues by a series of ultra-short laser pulses. However, the laser beam characteristics change dynamically due to interactions with birefringent corneal tissue, which may affect the outcome of the laser treatment. To better understand the effect the cornea has on the laser characteristics, we developed a system for measuring retardation and validated it with precise, standard phase retarders. Then we measured the phase retardation of FSLs through bovine corneas and found that there is a considerable, location dependent, variation in retardation values. This information can potentially help optimize FSL parameters to make their application in ophthalmic procedures safer and more effective.

Structure of corneal layers, collagen fibrils, and proteoglycans of tree shrew cornea

PubMed Central

Almubrad, Turki

2011-01-01

Purpose The stroma is the major part of the cornea, in which collagen fibrils and proteoglycans are distributed uniformly. We describe the ultrastructure of corneal layers, collagen fibrils (CF), and proteoglycans (PGs) in the tree shrew cornea. Methods Tree shrew corneas (5, 6, and 10 week old animals) and normal human corneas (24, 25, and 54 years old) were fixed in 2.5% glutaraldehyde containing cuprolinic blue in a sodium acetate buffer. The tissue was processed for electron microscopy. The ‘iTEM Olympus Soft Imaging Solutions GmbH’ program was used to measure the corneal layers, collagen fibril diameters and proteoglycan areas. Results The tree shrew cornea consists of 5 layers: the epithelium, Bowman’s layer, stroma, Descemet’s membrane, and endothelium. The epithelium was composed of squamous cells, wing cells and basal cells. The Bowman’s layer was 5.5±1.0 µm thick and very similar to a normal human Bowman’s layer. The stroma was 258±7.00 µm thick and consisted of collagen fibril lamellae. The lamellae were interlaced with one another in the anterior stroma, but ran parallel to one another in the middle and posterior stroma. Collagen fibrils were decorated with proteoglycan filaments with an area size of 390 ±438 nm2. The collagen fibril had a minimum diameter of 39±4.25 nm. The interfibrillar spacing was 52.91±6.07 nm. Within the collagen fibrils, very small electron-dense particles were present. Conclusions The structure of the tree shrew cornea is very similar to that of the normal human cornea. As is the case with the human cornea, the tree shrew cornea had a Bowman's layer, lamellar interlacing in the anterior stroma and electron-dense particles within the collagen fibrils. The similarities of the tree shrew cornea with the human cornea suggest that it could be a good structural model to use when studying changes in collagen fibrils and proteoglycans in non-genetic corneal diseases, such as ectasia caused after LASIK (laser-assisted in situ keratomileusis). PMID:21921979

A comparison of glycosaminoglycan distributions, keratan sulphate sulphation patterns and collagen fibril architecture from central to peripheral regions of the bovine cornea

PubMed Central

Ho, Leona T.Y.; Harris, Anthony M.; Tanioka, Hidetoshi; Yagi, Naoto; Kinoshita, Shigeru; Caterson, Bruce; Quantock, Andrew J.; Young, Robert D.; Meek, Keith M.

2014-01-01

This study investigated changes in collagen fibril architecture and the sulphation status of keratan sulphate (KS) glycosaminoglycan (GAG) epitopes from central to peripheral corneal regions. Freshly excised adult bovine corneal tissue was examined as a function of radial position from the centre of the cornea outwards. Corneal thickness, tissue hydration, hydroxyproline content, and the total amount of sulphated GAG were all measured. High and low-sulphated epitopes of keratan sulphate were studied by immunohistochemistry and quantified by ELISA. Chondroitin sulphate (CS) and dermatan sulphate (DS) distributions were observed by immunohistochemistry following specific enzyme digestions. Electron microscopy and X-ray fibre diffraction were used to ascertain collagen fibril architecture. The bovine cornea was 1021 ± 5.42 μm thick at its outer periphery, defined as 9–12 mm from the corneal centre, compared to 844 ± 8.10 μm at the centre. The outer periphery of the cornea was marginally, but not significantly, more hydrated than the centre (H = 4.3 vs. H = 3.7), and was more abundant in hydroxyproline (0.12 vs. 0.06 mg/mg dry weight of cornea). DMMB assays indicated no change in the total amount of sulphated GAG across the cornea. Immunohistochemistry revealed the presence of both high- and low-sulphated epitopes of KS, as well as DS, throughout the cornea, and CS only in the peripheral cornea before the limbus. Quantification by ELISA, disclosed that although both high- and low-sulphated KS remained constant throughout stromal depth at different radial positions, high-sulphated epitopes remained constant from the corneal centre to outer-periphery, whereas low-sulphated epitopes increased significantly. Both small angle X-ray diffraction and TEM analysis revealed that collagen fibril diameter remained relatively constant until the outer periphery was reached, after which fibrils became more widely spaced (from small angle x-ray diffraction analysis) and of larger diameter as they approached the sclera. Depth-profiled synchrotron microbeam analyses showed that, at different radial positions from the corneal centre outwards, fibril diameter was greater superficially than in deeper stromal regions. The interfibrillar spacing was also higher at mid-depth in the stroma than it was in anterior and posterior stromal regions. Collagen fibrils in the bovine cornea exhibited a fairly consistent spacing and diameter from the corneal centre to the 12 mm radial position, after which a significant increase was seen. While the constancy of the overall sulphation levels of proteoglycans in the cornea may correlate with the fibrillar architecture, there was no correlation between the latter and the distribution of low-sulphated KS. PMID:25019467

Prevalidation of a human cornea construct as an alternative to animal corneas for in vitro drug absorption studies.

PubMed

Hahne, Matthias; Zorn-Kruppa, Michaela; Guzman, Gustavo; Brandner, Johanna M; Haltner-Ukomado, Eleonore; Wätzig, Hermann; Reichl, Stephan

2012-08-01

The use of ophthalmic drugs has increased consistently over the past few decades. Currently, most research is conducted using in vivo and ex vivo animal experiments; however, they have many disadvantages, including ethical concerns, high costs, the questionable extension of animal results to humans, and poor standardization. Although several cell culture-based cornea models have been developed, none have been validated and accepted for general use. In this study, a standardized, three-dimensional model of the human cornea (Hemicornea, HC) based on immortalized human corneal cells and cultivated in serum-free conditions was developed for drug absorption studies and prevalidated using compounds with a wide range of molecular characteristics (sodium fluorescein, rhodamine B, fluorescein isothiocyanate-labeled dextran, aciclovir, bimatoprost, dexamethasone, and timolol maleate). The HC model was independently cultured in three different laboratories, and the intralaboratory and interlaboratory reproducibility was analyzed and compared with the rabbit cornea. This analysis showed that the HC has a barrier in the same range as excised animal corneas, although with a higher reproducibility and lower variability. Because of the demonstrated transferability, the HC represents a promising in vitro alternative to the use of ex vivo tissue and offers a well-defined and standardized system for drug absorption studies. Copyright © 2012 Wiley Periodicals, Inc.

Biomedical Research, Development, and Engineering at the Johns Hopkins University Applied Physics Laboratory Annual Report, October 1, 1975-September 30, 1976

DTIC Science & Technology

1976-09-01

Holographic Stress Testing of the surgicai. cornea, B. F. Hochheimer (APL) and J. .. Calkins (JHMI) §37 Constant Pressure Tonography, L. J. Viernatein...September 1976 HOLOGRAPHIC STRESS TESTING OF SUMMARY AND DISCUSSION THE SURGICAL CORNEA We have successfully demonstrated that holographic interferometry is...recording me- stress testing of corneal wounds in several thodology suitable for performing do sle-pulsed postoperative patients. The only stress used

The effect of single and repeated UVB radiation on rabbit cornea.

PubMed

Fris, Miroslav; Tessem, May-Britt; Cejková, Jitka; Midelfart, Anna

2006-12-01

Cumulative effect of ultraviolet radiation (UVR) is an important aspect of UV corneal damage. The purpose of this study was to apply high resolution magic angle spinning proton nuclear magnetic resonance (HR-MAS 1H NMR) spectroscopy to evaluate the effect of single and repeated UV radiation exposure of the same overall dose on the rabbit cornea. Corneal surfaces of 24 normal rabbit eyes were examined for the effects of UVB exposure (312 nm). In the first group (UVB1), animals were irradiated with a single dose (3.12 J/cm2; 21 min) of UVB radiation. The animals in the second group (UVB2) were irradiated three times for 7 min every other day (dose of 1.04 J/cm2; days 1, 3, 5) to give the same overall dose (3.12 J/cm2). The third group served as an untreated control group. One day after the last irradiation, the animals were sacrificed, and the corneas were removed and frozen. HR-MAS 1H NMR spectra from intact corneas were obtained. Special grouping patterns among the tissue samples and the relative percentage changes in particular metabolite concentrations were evaluated using modern statistical methods (multivariate analysis, one-way ANOVA). The metabolic profile of both groups of UVB-irradiated samples was significantly different from the control corneas. Substantial decreases in taurine, hypo-taurine and choline-derivatives concentrations and substantial elevation in glucose and betaine levels were observed following the UVR exposure. There was no significant difference between the effect of a single and repeated UVB irradiation of the same overall dose. For the first time, the effects of single and repeated UVR doses on the metabolic profile of the rabbit cornea were analysed and compared. The combination of HR-MAS 1H NMR spectroscopy and modern statistical methods (multivariate analysis, one-way ANOVA) proved suitable to assess the overall view of the metabolic alterations in the rabbit corneal tissue following UVB radiation exposure.

Hyaluronic acid coated poly-epsilon-caprolactone nanospheres deliver high concentrations of cyclosporine A into the cornea.

PubMed

Yenice, Irem; Mocan, Mehmet C; Palaska, Erhan; Bochot, Amélie; Bilensoy, Erem; Vural, Imran; Irkeç, Murat; Hincal, A Atilla

2008-09-01

The objective of this study was to determine cyclosporine A (Cy A) levels in ocular tissues and fluids after topical administration of poly-epsilon-caprolactone (PCL)/benzalkonium chloride (BKC) nanospheres and hyaluronic acid (HA) coated PCL/BKC nanospheres onto healthy rabbit corneas. Nanospheres were prepared by nanoprecipitation and purified by gradient-rate centrifugation. Cy A (0.1%) in either castor oil solution (group 1), PCL/BKC nanosphere formulation (group 2) or HA coated PCL/BKC nanosphere formulation (group 3) was instilled onto rabbit corneas. Tear samples were adsorbed onto Schirmer tear strips. Cy A concentrations of fluid (blood, aqueous humor, tear) and specimen extracts (cornea, conjunctiva, iris/ciliary body) were determined by high performance liquid chromatography-mass spectrometry (LC-MS). The mean corneal Cy A concentration obtained at 0.5, 1, 2, 4, 8 and 24h following instillation of the formulations ranged between 0.12 and 1.2 ng/mg tissue for group 1, 5.9-15.5 ng/mg tissue for group 2 and 11.4-23.0 ng/mg for group 3 (one-way analysis of variance (ANOVA) and pairwise tests (SNK (Student-Newman-Keuls) and Tukey); p<0.05). Conjunctival Cy A levels of group 2 and 3 were not significantly different at any of the time points tested. However, there was a significant difference between Cy A concentration of castor oil formulation and that of PCL/BKC nanosphere formulation at 1 and 8h (p<0.05). The mean iris/ciliary body concentrations obtained with the three formulations were not significantly different at any time point with the exception of group 2 levels being higher than those of groups 1 and 3 at 1h (p<0.05). The lowest ocular tear Cy A concentrations (16-114 ng/ml) were found following the instillation of HA coated PCL/BKC nanoparticles (group 3) during the time period tested. Cy A loaded PCL/BKC and HA coated PCL/BKC nanospheres are able to achieve high levels of Cy A in the cornea that is 10-15-fold higher than that is achieved with Cy A solution in castor oil. Nanosphere formulation and HA may play an important role in delivering high levels of cyclosporine A into the cornea.

Assessment of corneal hydration sensing in the terahertz band: in vivo results at 100 GHz

PubMed Central

Taylor, Zachary; Tewari, Pria; Sung, Shijun; Maccabi, Ashkan; Singh, Rahul; Culjat, Martin; Grundfest, Warren; Hubschman, Jean-Pierre; Brown, Elliott

2012-01-01

Abstract. Terahertz corneal hydration sensing has shown promise in ophthalmology applications and was recently shown to be capable of detecting water concentration changes of about two parts in a thousand in ex vivo corneal tissues. This technology may be effective in patient monitoring during refractive surgery and for early diagnosis and treatment monitoring in diseases of the cornea. In this work, Fuchs dystrophy, cornea transplant rejection, and keratoconus are discussed, and a hydration sensitivity of about one part in a hundred is predicted to be needed to successfully distinguish between diseased and healthy tissues in these applications. Stratified models of corneal tissue reflectivity are developed and validated using ex vivo spectroscopy of harvested porcine corneas that are hydrated using polyethylene glycol solutions. Simulation of the cornea’s depth-dependent hydration profile, from 0.01 to 100 THz, identifies a peak in intrinsic reflectivity contrast for sensing at 100 GHz. A 100 GHz hydration sensing system is evaluated alongside the current standard ultrasound pachymetry technique to measure corneal hydration in vivo in four rabbits. A hydration sensitivity, of three parts per thousand or better, was measured in all four rabbits under study. This work presents the first in vivo demonstration of remote corneal hydration sensing. PMID:23085925

Investigation of the formation mechanism and morphology of the features created in the interior of cornea by femtosecond laser pulses

NASA Astrophysics Data System (ADS)

Guo, Yizang; Vukelic, Sinisa

2015-03-01

Laser assisted corneal surgeries often rely on the nonlinear absorption effect of ultrafast lasers to induce features in the interior of the cornea without affecting the surface. In particular, corneal flap formation in femtosecond assisted Laser- Assisted in situ Keratomileusis (LASIK) is based on the bubble creation. This study focuses on the interaction between the tissue and the femtosecond laser. Interior of cornea is treated with tightly focused femtosecond laser pulses. Due to the nature of the process, heating of the tissue within and around the focal volume is practically instantaneous. The affected region is subject to thermoelastic stress that arises with the steep temperature elevation. To predict the size of the region subject to the morphological changes due to the laser treatment, the temperature field is calculated. Cavitation bubble initiation and expansion process, which acts as precursor to the stress induced tissue trauma, is studied as well. Theoretical findings are compared against experimental results. High-speed camera is utilized to assess the laser treatment process, showing the temporal development of the cavitation bubbles. The results obtained in this study facilitate a better understanding of the effects of femtosecond laser assisted corneal surgeries and help in choosing optimal laser parameters.

Experimental and model analysis on the temperature dynamics during diode laser welding of the cornea.

PubMed

Rossi, Francesca; Pini, Roberto; Menabuoni, Luca

2007-01-01

Corneal laser welding is a technique used clinically to induce the immediate sealing of corneal wounds. We present an experimental and model analysis of the temperature dynamics during diode laser-induced corneal welding, which is aimed at characterizing the mechanism of tissue fusion. Ex vivo tests were performed on porcine eyes in the typical irradiation conditions used for laser-induced suturing in cornea transplant. Three laser power densities (12.5 W/cm(2), 16.7 W/cm(2), 20.8 W/cm(2)) were tested. The superficial temperature of the cornea was measured by means of an infrared thermocamera. Experimental data were compared with the results of a three-dimensional (3D) model of a laser-welding process in the cornea, solved by the use of the Finite Element Method (FEM). The model solution and experimental results showed good agreement. The model was thus used to estimate the temperature enhancement inside the corneal wound and to calculate the thermal damage inside the tissue. The results indicated a selective, spatially confined heating effect that occurred at operative temperatures (59 to 66 degrees C) close to intermediate denaturation points of the stromal collagen, before its complete disorganization. No significant heat damage to the region of the laser-treated wound was evidenced in the operative irradiation conditions of corneal welding.

Dynamic behavior of photoablation products of corneal tissue in the mid-IR: a study with FELIX

NASA Astrophysics Data System (ADS)

Auerhammer, J. M.; Walker, R.; van der Meer, A. F. G.; Jean, B.

The properties of pulsed IR-laser ablation of biological soft tissue (porcine cornea) were studied in vitro systematically and quantitatively with a free-electron laser in the wavelength range 6<=λ<=20 μm at fluences ranging from 3.1 to 9.4 J/cm2. Dynamic parameters such as the extension of the ablation cloud, the initial velocity and momentum of the ablated particles as well as the ablation threshold, the ablated mass, and the particle size were investigated. The ablation plume was made visible with a stroboscopic technique. For a fluence of 3.1 J/cm2 the average initial velocity of the ejected particles was deduced from the extension of the plume to range from 120-400 m/s. Measurements of the recoil momentum using a sensitive pendulum led to values between 0.5 and 2.0 mmg/s. All measured properties were related to the spectroscopically determined absorption coefficient of cornea αcornea. Where absorption due to proteins is high (at λ=6.2 and 6.5 μm), ablated mass, velocity and recoil momentum behave according to αcornea. For the first time, variations of the ablation plume from pulse to pulse were observed. Those, as well as the particle size, not only depend on the absorption coefficient, but also on the predominant absorber.

Absence of beta-amyloid in cortical cataracts of donors with and without Alzheimer's disease.

PubMed

Michael, Ralph; Rosandić, Jurja; Montenegro, Gustavo A; Lobato, Elvira; Tresserra, Francisco; Barraquer, Rafael I; Vrensen, Gijs F J M

2013-01-01

Eye lenses from human donors with and without Alzheimer's disease (AD) were studied to evaluate the presence of amyloid in cortical cataract. We obtained 39 lenses from 21 postmortem donors with AD and 15 lenses from age-matched controls provided by the Banco de Ojos para Tratamientos de la Ceguera (Barcelona, Spain). For 17 donors, AD was clinically diagnosed by general physicians and for 4 donors the AD diagnosis was neuropathologically confirmed. Of the 21 donors with AD, 6 had pronounced bilateral cortical lens opacities and 15 only minor or no cortical opacities. As controls, 7 donors with pronounced cortical opacities and 8 donors with almost transparent lenses were selected. All lenses were photographed in a dark field stereomicroscope. Histological sections were analyzed using a standard and a more sensitive Congo red protocol, thioflavin staining and beta-amyloid immunohistochemistry. Brain tissue from two donors, one with cerebral amyloid angiopathy and another with advanced AD-related changes and one cornea with lattice dystrophy were used as positive controls for the staining techniques. Thioflavin, standard and modified Congo red staining were positive in the control brain tissues and in the dystrophic cornea. Beta-amyloid immunohistochemistry was positive in the brain tissues but not in the cornea sample. Lenses from control and AD donors were, without exception, negative after Congo red, thioflavin, and beta-amyloid immunohistochemical staining. The results of the positive control tissues correspond well with known observations in AD, amyloid angiopathy and corneas with lattice dystrophy. The absence of staining in AD and control lenses with the techniques employed lead us to conclude that there is no beta-amyloid in lenses from donors with AD or in control cortical cataracts. The inconsistency with previous studies of Goldstein et al. (2003) and Moncaster et al. (2010), both of which demonstrated positive Congo red, thioflavin, and beta-amyloid immunohistochemical staining in AD and Down syndrome lenses, is discussed. Copyright © 2012 Elsevier Ltd. All rights reserved.

Hydrogel Ring for Topical Drug Delivery to the Ocular Posterior Segment.

PubMed

Shikamura, Yuko; Yamazaki, Yoshiko; Matsunaga, Toru; Sato, Takao; Ohtori, Akira; Tojo, Kakuji

2016-05-01

To investigate the efficacy of a topical hydrogel ring for drug delivery to the posterior segment of the rabbit eye. Novel hydrogel corneal lenses (CL), scleral/corneal lenses (S/CL), and rings were prepared using poly(hydroxyethyl methacrylate). The devices were immersed in 0.3% ofloxacin ophthalmic solution (OOS) to homogeneously distribute the drug throughout the hydrogel. The medicated CL, S/CL, Ring 1 (standard ring), or Ring 2 (shape-optimized ring) was applied to the surface of the cornea, cornea/bulbar conjunctiva, or bulbar conjunctiva of albino rabbits, respectively. Medicated rings did not touch the corneal surface. In another group, one OOS drop was administered to the eye. After 0.25-8 hours, the hydrogel devices were removed and ocular tissues were harvested. High-performance liquid chromatography (HPLC) was used to measure the ofloxacin concentration in the devices and tissues. The drug concentrations in the posterior segment tissues were compared among ofloxacin delivery methods. One hour after placement, eyes treated with Ring 1 or S/CL had markedly higher ofloxacin levels in the posterior segment tissues (conjunctiva, sclera, and retina/choroid) than eyes treated with topical OOS or a CL. Lower levels of ofloxacin were found in anterior segment tissues (cornea and aqueous humor) in eyes treated with Ring 1 compared to those treated with S/CL. Ring 2 most effectively delivered ofloxacin to the retina/choroid. The tissue ofloxacin concentration in the fellow eye was markedly lower than the eye treated with Ring 2. Our results suggest that hydrogel rings are effective in delivering topical ophthalmic drugs to the posterior segment. The drugs are most likely delivered via the transconjunctival/scleral route by lateral diffusion across the bulbar conjunctiva and through the sclera. Systemic drug delivery to the posterior segment is minimal.

Determination of permeability coefficients of ophthalmic drugs through different layers of porcine, rabbit and bovine eyes.

PubMed

Loch, Christian; Zakelj, Simon; Kristl, Albin; Nagel, Stefan; Guthoff, Rudolf; Weitschies, Werner; Seidlitz, Anne

2012-08-30

To treat ophthalmic diseases like glaucoma or inflammatory disorders topically applied ophthalmic formulations such as eye drops are usually used. In addition, novel ophthalmic implants releasing drug substances locally into different parts of the eye are available today. In the work presented here, the permeability coefficients of selected drugs (ciprofloxacin hydrochloride, lidocaine hydrochloride, timolol maleate) for ophthalmic tissues were determined using side-by-side diffusion chambers (so-called Ussing chambers). Sclera, conjunctiva, cornea, choroidea-retina-complex and a complex of conjunctiva-sclera-choroidea-retina were excised from fresh porcine, rabbit and bovine eyes. In the porcine eye tissues the highest P(app) values were obtained for conjunctiva with the exception of lidocaine. Therefore, it can be estimated that a certain amount of drug diffuses or is transported through conjunctiva after application. The P(app) values for sclera were also higher than those for cornea and even more, the surface area of sclera which is available for drug absorption is much larger than that of cornea when applying an implant. The obtained permeability coefficients for sclera and conjunctiva indicate that the administration of periocular implants can be an alternative to topically applied formulations. The complexes of the tissues were a significantly (p<0.01) stronger barrier to the investigated substances than the separated tissues. Distinct differences in permeability coefficients between the investigated animal tissues were observed. Overall the highest P(app) values for all mounted tissues were obtained with the rabbit, followed by porcine and bovine eyes. Because of these distinct interspecies differences one must be very careful when selecting the proper animal model for the permeability experiments. Copyright © 2012 Elsevier B.V. All rights reserved.

The New Zealand National Eye Bank study: trends in the acquisition and storage of corneal tissue over the decade 2000 to 2009.

PubMed

Cunningham, William J; Moffatt, S Louise; Brookes, Nigel H; Twohill, Helen C; Pendergrast, David G C; Stewart, Joanna M; McGhee, Charles N J

2012-05-01

To evaluate trends in the acquisition, storage, and utilization of donated corneal tissue in New Zealand, 2000 to 2009. The New Zealand National Eye Bank records were analyzed for the decade January 2000 to December 2009. Variables analyzed included donor demographics (age, sex, and ethnicity), donor source, donor cause of death, death-to-preservation interval (DPI), corneal storage time, tissue contamination, endothelial assessment, cornea suitability for transplantation, and corneal tissue utilization. A total of 1268 eye donors were identified during the 10-year period. Overall, 36% (n = 457) were female and 64% male (n = 813). Median donor age was 67 years, and 23% of donors were younger than 50 years (range, 5-90 years). There was a decrease in donor age over the decade (P = 0.006). The median DPI was 18.5 hours. No relationship was identified between cornea suitability for transplantation and DPI (P = 0.28) or donor gender (P = 0.54). There was a low microbial contamination rate (1%). Human immunodeficiency virus, hepatitis B, or hepatitis C serology was positive in 48 donors (4%). Overall, 90% of corneas were suitable for transplantation with a high utilization rate (88%). A novel association was identified between male sex and lower corneal endothelial cell density (P = 0.03). This New Zealand National Eye Bank analysis identified trends in the acquisition, storage, and utilization of donated corneal tissue throughout New Zealand over the past decade and provides valuable additional information to the international eye bank data.

High resolution macroscopy (HRMac) of the eye using nonlinear optical imaging

NASA Astrophysics Data System (ADS)

Winkler, Moritz; Jester, Bryan E.; Nien-Shy, Chyong; Chai, Dongyul; Brown, Donald J.; Jester, James V.

2010-02-01

Non-linear optical (NLO) imaging using femtosecond lasers provides a non-invasive means of imaging the structural organization of the eye through the generation of second harmonic signals (SHG). While NLO imaging is able to detect collagen, the small field of view (FoV) limits the ability to study how collagen is structurally organized throughout the larger tissue. To address this issue we have used computed tomography on optical and mechanical sectioned tissue to greatly expand the FoV and provide high resolution macroscopic (HRMac) images that cover the entire tissue (cornea and optic nerve head). Whole, fixed cornea (13 mm diameter) or optic nerve (3 mm diameter) were excised and either 1) embedded in agar and sectioned using a vibratome (200-300 um), or 2) embedded in LR White plastic resin and serially sectioned (2 um). Vibratome and plastic sections were then imaged using a Zeiss LSM 510 Meta and Chameleon femtosecond laser to generate NLO signals and assemble large macroscopic 3-dimensional tomographs with high resolution that varied in size from 9 to 90 Meg pixels per plane having a resolution of 0.88 um lateral and 2.0 um axial. 3-D reconstructions allowed for regional measurements within the cornea and optic nerve to quantify collagen content, orientation and organization over the entire tissue. We conclude that NLO based tomography to generate HRMac images provides a powerful new tool to assess collagen structural organization. Biomechanical testing combined with NLO tomography may provide new insights into the relationship between the extracellular matrix and tissue mechanics.

Proteolytic processing of connective tissue growth factor in normal ocular tissues and during corneal wound healing.

PubMed

Robinson, Paulette M; Smith, Tyler S; Patel, Dilan; Dave, Meera; Lewin, Alfred S; Pi, Liya; Scott, Edward W; Tuli, Sonal S; Schultz, Gregory S

2012-12-13

Connective tissue growth factor (CTGF) is a fibrogenic cytokine that is up-regulated by TGF-β and mediates most key fibrotic actions of TGF-β, including stimulation of synthesis of extracellular matrix and differentiation of fibroblasts into myofibroblasts. This study addresses the role of proteolytic processing of CTGF in human corneal fibroblasts (HCF) stimulated with TGF-β, normal ocular tissues and wounded corneas. Proteolytic processing of CTGF in HCF cultures, normal animal eyes, and excimer laser wounded rat corneas were examined by Western blot. The identity of a 21-kDa band was determined by tandem mass spectrometry, and possible alternative splice variants of CTGF were assessed by 5' Rapid Amplification of cDNA Ends (RACE). HCF stimulated by TGF-β contained full length 38-kDa CTGF and fragments of 25, 21, 18, and 13 kDa, while conditioned medium contained full length 38- and a 21-kDa fragment of CTGF that contained the middle "hinge" region of CTGF. Fragmentation of recombinant CTGF incubated in HCF extracts was blocked by the aspartate protease inhibitor, pepstatin. Normal mouse, rat, and rabbit whole eyes and rabbit ocular tissues contained abundant amounts of C-terminal 25- and 21-kDa fragments and trace amounts of 38-kDa CTGF, although no alternative transcripts were detected. All forms of CTGF (38, 25, and 21 kDa) were detected during healing of excimer ablated rat corneas, peaking on day 11. Proteolytic processing of 38-kDa CTGF occurs during corneal wound healing, which may have important implications in regulation of corneal scar formation.

Design and evaluation of artificial cornea with core-skirt design using polyhydroxyethyl methacrylate and graphite.

PubMed

Sinha, Mukty; Gupte, Tanvi

2017-06-10

Artificial cornea is the effective treatment option for corneal blindness. One of the challenges with the artificial cornea is limited, or no tissue integration necessitates reimplantation due to necrosis or corneal melting. We propose here a new formulation approach for core-skirt incorporating graphite in the outer skirt region to improve cell adhesion. Hydroxyethyl methacrylate (HEMA) and ethylene glycol dimethacrylate were procured from Sigma-Aldrich. Polyhydroxyethyl methacrylate (PHEMA) was synthesized by free radical polymerization of HEMA. PHEMA hydrogel core with graphite incorporated skirt was developed with the help of mould and spacer. Pores were introduced into the skirt by salt leaching technique using sodium chloride as porogen. The porous skirt was improved for its aesthetic appeal of black colour and mechanical strength to sustain intraocular pressure by incorporating graphite. The material properties of the newly developed design were evaluated in terms of wetting behaviour, mechanical strength, water vapour permeability, degradation profile and cell adhesion. The polymerization of HEMA was confirmed by thin layer chromatography and FTIR. Water content of the polymeric film was optimized at 50% where maximum transparency with required refractive index of 1.4 was obtained. The concentration of salt vital for the essential porosity was also optimized using optical microscopy and scanning electron microscopy. Other properties, namely mechanical strength, water vapour transmission rate and degradation behaviour, showed that the developed design is suitable for ocular applications. Furthermore, cell adhesion study confirmed tissue adhesion in the skirt region but absent in the core. The core-skirt design may offer an efficient cornea replacement alternative with enhanced tissue integration in addition to desired mechanical behaviour with a clear and aesthetic vision.

Low-Level Blast Exposure Increases Transient Receptor Potential Vanilloid 1 (TRPV1) Expression in the Rat Cornea

PubMed Central

Por, Elaine D.; Choi, Jae-Hyek; Lund, Brian J.

2016-01-01

ABSTRACT Background: Blast-related ocular injuries sustained by military personnel have led to rigorous efforts to elucidate the effects of blast exposure on neurosensory function. Recent studies have provided some insight into cognitive and visual deficits sustained following blast exposure; however, limited data are available on the effects of blast on pain and inflammatory processes. Investigation of these secondary effects of blast exposure is necessary to fully comprehend the complex pathophysiology of blast-related injuries. The overall purpose of this study is to determine the effects of single and repeated blast exposure on pain and inflammatory mediators in ocular tissues. Methods: A compressed air shock tube was used to deliver a single or repeated blast (68.0 ± 2.7 kPa) to anesthetized rats daily for 5 days. Immunohistochemistry was performed on ocular tissues to determine the expression of the transient receptor potential vanilloid 1 (TRPV1) channel, calcitonin gene-related peptide (CGRP), substance P (SP), and endothelin-1 (ET-1) following single and repeated blast exposure. Neutrophil infiltration and myeloperoxidase (MPO) expression were also assessed in blast tissues via immunohistochemistry and enzyme-linked immunosorbent assay (ELISA) analysis, respectively. Results: TRPV1 expression was increased in rat corneas exposed to both single and repeated blast. Increased secretion of CGRP, SP, and ET-1 was also detected in rat corneas as compared to control. Moreover, repeated blast exposure resulted in neutrophil infiltration in the cornea and stromal layer as compared to control animals. Conclusion: Single and repeated blast exposure resulted in increased expression of TRPV1, CGRP, SP, and ET-1 as well as neutrophil infiltration. Collectively, these findings provide novel insight into the activation of pain and inflammation signaling mediators following blast exposure. PMID:27049881

DNA methyl transferases are differentially expressed in the human anterior eye segment.

PubMed

Bonnin, Nicolas; Belville, Corinne; Chiambaretta, Frédéric; Sapin, Vincent; Blanchon, Loïc

2014-08-01

DNA methylation is an epigenetic mark involved in the control of genes expression. Abnormal epigenetic events have been reported in human pathologies but weakly documented in eye diseases. The purpose of this study was to establish DNMT mRNA and protein expression levels in the anterior eye segment tissues and their related (primary or immortalized) cell cultures as a first step towards future in vivo and in vitro methylomic studies. Total mRNA was extracted from human cornea, conjunctiva, anterior lens capsule, trabeculum and related cell cultures (cornea epithelial, trabecular meshwork, keratocytes for primary cells; and HCE, Chang, B-3 for immortalized cells). cDNA was quantified by real-time PCR using specific primers for DNMT1, 2, 3A, 3B and 3L. Immunolocalization assays were carried out on human cornea using specific primary antibodies for DNMT1, 2 and 3A, 3B and 3L. All DNMT transcripts were detected in human cornea, conjunctiva, anterior lens capsule, trabeculum and related cells but showed statistically different expression patterns between tissues and cells. DNMT2 protein presented a specific and singular expression pattern in corneal endothelium. This study produced the first inventory of the expression patterns of DNMTs in human adult anterior eye segment. Our research highlights that DNA methylation cannot be ruled out as a way to bring new insights into well-known ocular diseases. In addition, future DNA methylation studies using various cells as experimental models need to be conducted with attention to approach the results analysis from a global tissue perspective. © 2014 Acta Ophthalmologica Scandinavica Foundation. Published by John Wiley & Sons Ltd.

Optical System Design for Noncontact, Normal Incidence, THz Imaging of in vivo Human Cornea.

PubMed

Sung, Shijun; Dabironezare, Shahab; Llombart, Nuria; Selvin, Skyler; Bajwa, Neha; Chantra, Somporn; Nowroozi, Bryan; Garritano, James; Goell, Jacob; Li, Alex; Deng, Sophie X; Brown, Elliott; Grundfest, Warren S; Taylor, Zachary D

2018-01-01

Reflection mode Terahertz (THz) imaging of corneal tissue water content (CTWC) is a proposed method for early, accurate detection and study of corneal diseases. Despite promising results from ex vivo and in vivo cornea studies, interpretation of the reflectivity data is confounded by the contact between corneal tissue and dielectric windows used to flatten the imaging field. Herein, we present an optical design for non-contact THz imaging of cornea. A beam scanning methodology performs angular, normal incidence sweeps of a focused beam over the corneal surface while keeping the source, detector, and patient stationary. A quasioptical analysis method is developed to analyze the theoretical resolution and imaging field intensity profile. These results are compared to the electric field distribution computed with a physical optics analysis code. Imaging experiments validate the optical theories behind the design and suggest that quasioptical methods are sufficient for designing of THz corneal imaging systems. Successful imaging operations support the feasibility of non-contact in vivo imaging. We believe that this optical system design will enable the first, clinically relevant, in vivo exploration of CTWC using THz technology.

Corneal NF-kappaB activity is necessary for the retention of transparency in the cornea of UV-B-exposed transgenic reporter mice.

PubMed

Alexander, George; Carlsen, Harald; Blomhoff, Rune

2006-04-01

To determine the dynamics of Nuclear Factor-kappaB (NF-kappaB) in murine corneal pathology and the role of NF-kappaB in maintaining corneal clarity after ultraviolet B radiation insult, transgenic mice containing NF-kappaB-luciferase reporter were exposed to LPS (bacterial lipopolysaccharide), TNF-alpha (Tumor Necrosis Factor-alpha) or 4 kJ m(-2) UV-B radiation. NF-kappaB decoy oligonucleotides were also administered in some of the UV-B experiments. Following various exposure times, the mice were sacrificed and whole eyes or corneal tissues were obtained. Whole eyes were examined for scattering using a point-source optical imaging technique. Tissue homogenates were examined for luciferase activity using a luminometer. TNF-alpha and LPS-injected NF-kappaB-luciferase transgenic mice demonstrated 3-10-fold increases in cornea NF-kappaB with peak activities at 4 and 6 hr post-injection, respectively. Mice exposed to 4 kJ m(-2) UV-B exhibited a 3-fold increase in NF-kappaB activity 4 hr post-exposure. The administration of NF-kappaB-decoy oligonucleotides to mice had the effect of reducing UV-B-induced NF-kappaB activity in the cornea and significantly increasing the amount of light scattering in UV-B exposed corneas 7 days post-UV-B exposure when compared to sham injected mice. These results indicate that NF-kappaB is activated in cornea in pathologies that involves increased plasma levels of LPS and TNF-alpha, as well as direct UV-B exposure, and suggest that NF-kappaB activation play an essential part in the corneal healing process.

Hypothermic preservation of corneas in a hyperkalaemic solution (CPTES): II. Extended storage in the presence of chondroitin sulphate.

PubMed Central

Taylor, M J; Hunt, C J; Madden, P W

1989-01-01

Periods of preservation for donor corneas, even for short times, are necessary to facilitate optimum conditions in penetrating keratoplasty. However, current techniques for corneal storage at low temperatures may not provide optimal conditions for maintaining tissue integrity. In particular, the ionic composition of the storage medium has received little attention since it has been assumed throughout that the normal complement of ions in tissue culture media will also be suitable for preservation at reduced temperatures. This study extends our previous investigations on the merits of using CPTES (corneal-potassium-TES), a potassium-rich balanced salt solution containing an impermeant anionic pH buffer (TES), as a storage solution specifically designed to prevent the loss of intracellular potassium and minimise endothelial cell swelling during the time that the normal regulatory processes are switched off. The effect of adding the natural polymer chondroitin sulphate (CS) as a colloid osmotic agent to the hyperkalaemic storage medium is now examined. Corneas stored in CPTES containing 2.5% chondroitin sulphate retained a very high level of structural and functional integrity after three, five, and seven days storage at 0 degrees C; furthermore, stromal swelling was restricted to only 21%. All corneas stored in CPTES + 2.5% CS showed active endothelial function by thinning efficiently at rates that were greater than those previously reported for rabbit corneas stored for similar lengths of time in either M-K medium or K-sol. The zwitterionic buffers TES and HEPES were interchangeable in the hyperkalaemic solution and were non-toxic to corneal endothelium at a concentration of 100 mM. These compounds offer excellent pH buffering in bicarbonate-free medium. Images PMID:2510816

Comparison of retina damage thresholds simulating the femtosecond-laser in situ keratomileusis (fs-LASIK) process with two laser systems in the CW- and fs-regime

NASA Astrophysics Data System (ADS)

Sander, M.; Minet, O.; Zabarylo, U.; Müller, M.; Tetz, M. R.

2012-04-01

The femtosecond-laser in situ keratomileusis procedure affords the opportunity to correct ametropia by cutting transparent corneal tissue with ultra-short laser pulses. Thereby the tissue cut is generated by a laser-induced optical breakdown in the cornea with ultra-short laser pulses in the near-infrared range. Compared to standard procedures such as photorefractive keratectomy and laser in-situ keratomileusis with the excimer laser, where the risk potential for the eye is low due to the complete absorption of ultraviolet irradiation from corneal tissue, only a certain amount of the pulse energy is deposited in the cornea during the fs-LASIK process. The remaining energy propagates through the eye and interacts with the retina and the strong absorbing tissue layers behind. The objective of the presented study was to determine and compare the retina damage thresholds during the fs-LASIK process simulated with two various laser systems in the CW- and fs-regime.

The potential role of bioengineering and three-dimensional printing in curing global corneal blindness.

PubMed

Ludwig, Parker E; Huff, Trevor J; Zuniga, Jorge M

2018-01-01

An insufficiency of accessible allograft tissue for corneal transplantation leaves many impaired by untreated corneal disease. There is promise in the field of regenerative medicine for the development of autologous corneal tissue grafts or collagen-based scaffolds. Another approach is to create a suitable corneal implant that meets the refractive needs of the cornea and is integrated into the surrounding tissue but does not attempt to perfectly mimic the native cornea on a cellular level. Materials that have been investigated for use in the latter concept include natural polymers such as gelatin, semisynthetic polymers like gelatin methacrylate, and synthetic polymers. There are advantages and disadvantages inherent in natural and synthetic polymers: natural polymers are generally more biodegradable and biocompatible, while synthetic polymers typically provide greater control over the characteristics or property adjustment of the materials. Additive manufacturing could aid in the precision production of keratoprostheses and the personalization of implants.

The potential role of bioengineering and three-dimensional printing in curing global corneal blindness

PubMed Central

Ludwig, Parker E; Huff, Trevor J; Zuniga, Jorge M

2018-01-01

An insufficiency of accessible allograft tissue for corneal transplantation leaves many impaired by untreated corneal disease. There is promise in the field of regenerative medicine for the development of autologous corneal tissue grafts or collagen-based scaffolds. Another approach is to create a suitable corneal implant that meets the refractive needs of the cornea and is integrated into the surrounding tissue but does not attempt to perfectly mimic the native cornea on a cellular level. Materials that have been investigated for use in the latter concept include natural polymers such as gelatin, semisynthetic polymers like gelatin methacrylate, and synthetic polymers. There are advantages and disadvantages inherent in natural and synthetic polymers: natural polymers are generally more biodegradable and biocompatible, while synthetic polymers typically provide greater control over the characteristics or property adjustment of the materials. Additive manufacturing could aid in the precision production of keratoprostheses and the personalization of implants. PMID:29686829

Interplay of wavelength, fluence and spot-size in free-electron laser ablation of cornea.

PubMed

Hutson, M Shane; Ivanov, Borislav; Jayasinghe, Aroshan; Adunas, Gilma; Xiao, Yaowu; Guo, Mingsheng; Kozub, John

2009-06-08

Infrared free-electron lasers ablate tissue with high efficiency and low collateral damage when tuned to the 6-microm range. This wavelength-dependence has been hypothesized to arise from a multi-step process following differential absorption by tissue water and proteins. Here, we test this hypothesis at wavelengths for which cornea has matching overall absorption, but drastically different differential absorption. We measure etch depth, collateral damage and plume images and find that the hypothesis is not confirmed. We do find larger etch depths for larger spot sizes--an effect that can lead to an apparent wavelength dependence. Plume imaging at several wavelengths and spot sizes suggests that this effect is due to increased post-pulse ablation at larger spots.

Pretreatment methods to improve nerve immunostaining in corneas from long-term fixed embryonic quail eyes

NASA Technical Reports Server (NTRS)

Barrett, J. E.; Wells, D. C.; Conrad, G. W.

1999-01-01

Pretreatment methods were used to improve neurofilament immunostaining in corneas from embryonic day 16 Japanese quail corneas that had been stored in fixative solution for several months. A sequential combination of the following three pretreatments: brief microwave heating in saline, followed by extraction with sodium dodecyl sulfate (SDS) at 37 degrees C, followed by digestion with hyaluronidase at 37 degrees C, produced significantly increased antibody staining of corneal neurofilament proteins, compared with embryonic corneas subjected to no prior pretreatments or to single or two-step protocols. After applying the sequence of all three pretreatments, darkest nerve staining and increased numbers of fine branches were observed, together with lower background staining. Thus, the result of applying the three-step pretreatment sequence is better than that of applying any of its component single pretreatments or even combinations of any two of them. These findings therefore suggest that each of these three pretreatments causes a unique effect, beneficial to immunostaining of neurofilament proteins, and that their individual effects are independent and additive. In addition to embryonic corneas, the three-step procedure also may be useful for immunostaining of nerves in other very delicate, highly-hydrated tissues containing an abundance of extracellular matrix.

Healing of corneal epithelial wounds in marine and freshwater fish.

PubMed

Ubels, J L; Edelhauser, H F

The corneal epithelium of a fish is in direct contact with the aquatic environment and is a barrier to movement of ions and water into and through the cornea. This tissue layer is thus important in maintenance of corneal transparency. When the epithelium is wounded, its protective function is lost and corneal transparency remains compromised until the epithelial barrier is re-established. This study was undertaken to investigate the healing response of the fish cornea to epithelial abrasion. Wounds were stained with fluorescein and photographed during healing. Wound areas were measured by planimetry. The cornea of the sculpin, a marine teleost, becomes edematous after wounding and heals at 2.54 to 3.42 mm2/hr. Nonswelling corneas of the elasmobranchs--dogfish shark and skate--heal at 1.29 mm2/hr, respectively. The wounded eye of the rainbow trout, a freshwater teleost, is stressed by the low osmolality of the environment. Severe corneal edema and cataracts develop following epithelial wounding, and the cornea heals at 0.64 mm2/hr. Although the healing rates in teleosts differ from those in mammals, histology shows that the corneal healing mechanism is essentially the same in fish and mammals.

Fundus oculi pigmentation studies simulating the fs-LASIK process Fundus oculi pigmentation studies simulating the fs-LASIK process

NASA Astrophysics Data System (ADS)

Sander, M.; Minet, O.; Zabarylo, U.; Müller, M.; Tetz, M. R.

2012-06-01

The femtosecond-laser in situ keratomileusis (fs-LASIK) technique has successfully entered the refractive surgery market to correct ametropia by cutting transparent corneal tissue with ultra-short laser pulses based on photodisruption. The laser pulses in the near infrared range (NIR) generate a laser-induced breakdown (LIOB) in the cornea. By propagating through the eye, a certain amount of the pulse is deposited in the cornea and the remaining energy interacts with the strong absorbing tissue behind. Due to the absorption by the retinal pigment epithelium and the transfer of the thermal energy to surrounding tissue, the transmitted energy can induce damage to the retina. The aim of this project was to find out the threshold influences concerning the tissue and the correlation between the results of the macroscopical appraisal and the fundus oculi pigmentation by simulating the fs-LASIK procedure with two various laser systems in the continuous wave (CW) and fs-regime. Therefore ex-vivo determinations were carried out macroscopically and histopathologically on porcine tissue.

Ophthalmic applications of confocal microscopy: diagnostics, refractive surgery, and eye banking

NASA Astrophysics Data System (ADS)

Masters, Barry R.

1990-11-01

Confocal microscopy of ocular tissue provides two advantages over traditional imaging techniques: increased range and transverse resolution and increased contrast. The semitransparent cornea and ocular lens in the living eye can be optically sectioned and observed by reflected light confocal microscopy. Within the cornea we observed various cell components nerve fibers nerve cell bodies and fibrous networks. The confocal microscopic images from the in-situ ocular lens show the lens capsule the lens epithelium and the individual lens fibrils. All of the reflected light confocal microscopic images have high contrast and high resolution. Some of the applications of confocal imaging in ophthalmology include: diagnostics of the cornea and the ocular lens examination prior to and after refractive surgery examination of intraocular lenses (IOL) and examination of eye bank material. Other ophthalmic uses of confocal imaging include: studies of wound healing therapeutics and the effects of contact lenses on the cornea. The proposed features of a clinical confocal microscope are reviewed. 2.

Elastography methods applicable to the eye

NASA Astrophysics Data System (ADS)

Khan, Altaf A.; Cortina, Soledad M.; Chamon, Wallace; Royston, Thomas J.

2014-02-01

Elastography is the mapping of tissues and cells by their respective mechanical properties, such as elasticity and viscosity. Our interest primarily lies in the human eye. Combining Scanning Laser Doppler Vibrometry (SLDV) with geometrically focused mechanical vibratory excitations of the cornea, it is possible to reconstruct these mechanical properties of the cornea. Experiments were conducted on phantom corneas as well as excised donor human corneas to test feasibility and derive a method of modeling. Finite element analysis was used to recreate the phantom studies and corroborate with the experimental data. Results are in close agreement. To further expand the study, lamb eyes were used in MR Elastography studies. 3D wave reconstruction was created and elastography maps were obtained. With MR Elastography, it would be possible to noninvasively measure mechanical properties of anatomical features not visible to SLDV, such as the lens and retina. Future plans include creating a more robust finite element model, improving the SLDV method for in-vivo application, and continuing experiments with MR Elastography.

Collagen in Human Tissues: Structure, Function, and Biomedical Implications from a Tissue Engineering Perspective

NASA Astrophysics Data System (ADS)

Balasubramanian, Preethi; Prabhakaran, Molamma P.; Sireesha, Merum; Ramakrishna, Seeram

The extracellular matrix is a complex biological structure encoded with various proteins, among which the collagen family is the most significant and abundant of all, contributing 30-35% of the whole-body protein. "Collagen" is a generic term for proteins that forms a triple-helical structure with three polypeptide chains, and around 29 types of collagen have been identified up to now. Although most of the members of the collagen family form such supramolecular structures, extensive diversity exists between each type of collagen. The diversity is not only based on the molecular assembly and supramolecular structures of collagen types but is also observed within its tissue distribution, function, and pathology. Collagens possess complex hierarchical structures and are present in various forms such as collagen fibrils (1.5-3.5 nm wide), collagen fibers (50-70 nm wide), and collagen bundles (150-250 nm wide), with distinct properties characteristic of each tissue providing elasticity to skin, softness of the cartilage, stiffness of the bone and tendon, transparency of the cornea, opaqueness of the sclera, etc. There exists an exclusive relation between the structural features of collagen in human tissues (such as the collagen composition, collagen fibril length and diameter, collagen distribution, and collagen fiber orientation) and its tissue-specific mechanical properties. In bone, a transverse collagen fiber orientation prevails in regions of higher compressive stress whereas longitudinally oriented collagen fibers correlate to higher tensile stress. The immense versatility of collagen compels a thorough understanding of the collagen types and this review discusses the major types of collagen found in different human tissues, highlighting their tissue-specific uniqueness based on their structure and mechanical function. The changes in collagen during a specific tissue damage or injury are discussed further, focusing on the many tissue engineering applications for which collagen scaffolds are currently being applied.

Corneal stroma microfibrils.

PubMed

Hanlon, Samuel D; Behzad, Ali R; Sakai, Lynn Y; Burns, Alan R

2015-03-01

Elastic tissue was first described well over a hundred years ago and has since been identified in nearly every part of the body. In this review, we examine elastic tissue in the corneal stroma with some mention of other ocular structures which have been more thoroughly described in the past. True elastic fibers consist of an elastin core surrounded by fibrillin microfibrils. However, the presence of elastin fibers is not a requirement and some elastic tissue is comprised of non-elastin-containing bundles of microfibrils. Fibers containing a higher relative amount of elastin are associated with greater elasticity and those without elastin, with structural support. Recently it has been shown that the microfibrils, not only serve mechanical roles, but are also involved in cell signaling through force transduction and the release of TGF-β. A well characterized example of elastin-free microfibril bundles (EFMBs) is found in the ciliary zonules which suspend the crystalline lens in the eye. Through contraction of the ciliary muscle they exert enough force to reshape the lens and thereby change its focal point. It is believed that the molecules comprising these fibers do not turn-over and yet retain their tensile strength for the life of the animal. The mechanical properties of the cornea (strength, elasticity, resiliency) would suggest that EFMBs are present there as well. However, many authors have reported that, although present during embryonic and early postnatal development, EFMBs are generally not present in adults. Serial-block-face imaging with a scanning electron microscope enabled 3D reconstruction of elements in murine corneas. Among these elements were found fibers that formed an extensive network throughout the cornea. In single sections these fibers appeared as electron dense patches. Transmission electron microscopy provided additional detail of these patches and showed them to be composed of fibrils (∼10 nm diameter). Immunogold evidence clearly identified these fibrils as fibrillin EFMBs and EFMBs were also observed with TEM (without immunogold) in adult mammals of several species. Evidence of the presence of EFMBs in adult corneas will hopefully pique an interest in further studies that will ultimately improve our understanding of the cornea's biomechanical properties and its capacity to repair. Copyright © 2015 Elsevier Ltd. All rights reserved.

Corneal Stroma Microfibrils

PubMed Central

Hanlon, Samuel D.; Behzad, Ali R.; Sakai, Lynn Y.; Burns, Alan R.

2015-01-01

Elastic tissue was first described well over a hundred years ago and has since been identified in nearly every part of the body. In this review, we examine elastic tissue in the corneal stroma with some mention of other ocular structures which have been more thoroughly described in the past. True elastic fibers consist of an elastin core surrounded by fibrillin microfibrils. However, the presence of elastin fibers is not a requirement and some elastic tissue is comprised of non-elastin-containing bundles of microfibrils. Fibers containing a higher relative amount of elastin are associated with greater elasticity and those without elastin, with structural support. Recently it has been shown that the microfibrils, not only serve mechanical roles, but are also involved in cell signaling through force transduction and the release of TGF-β. A well characterized example of elastin-free microfibril bundles (EFMBs) is found in the ciliary zonules which suspend the crystalline lens in the eye. Through contraction of the ciliary muscle they exert enough force to reshape the lens and thereby change its focal point. It is believed that the molecules comprising these fibers do not turn-over and yet retain their tensile strength for the life of the animal. The mechanical properties of the cornea (strength, elasticity, resiliency) would suggest that EFMBs are present there as well. However, many authors have reported that, although present during embryonic and early postnatal development, EFMBs are generally not present in adults. Serial-block-face imaging with a scanning electron microscope enabled 3D reconstruction of elements in murine corneas. Among these elements were found fibers that formed an extensive network throughout the cornea. In single sections these fibers appeared as electron dense patches. Transmission electron microscopy provided additional detail of these patches and showed them to be composed of fibrils (∼10nm diameter). Immunogold evidence clearly identified these fibrils as fibrillin EFMBs and EFMBs were also observed with TEM (without immunogold) in adult mammals of several species. Evidence of the presence of EFMBs in adult corneas will hopefully pique an interest in further studies that will ultimately improve our understanding of the cornea's biomechanical properties and its capacity to repair. PMID:25613072

Ex vivo rabbit and human corneas as models for bacterial and fungal keratitis.

PubMed

Pinnock, Abigail; Shivshetty, Nagaveni; Roy, Sanhita; Rimmer, Stephen; Douglas, Ian; MacNeil, Sheila; Garg, Prashant

2017-02-01

In the study of microbial keratitis, in vivo animal models often require a large number of animals, and in vitro monolayer cell culture does not maintain the three-dimensional structure of the tissues or cell-to-cell communication of in vivo models. Here, we propose reproducible ex vivo models of single- and dual-infection keratitis as an alternative to in vivo and in vitro models. Excised rabbit and human corneoscleral rims maintained in organ culture were infected using 10 8 cells of Staphylococcus aureus, Pseudomonas aeruginosa, Candida albicans or Fusarium solani. The infection was introduced by wounding with a scalpel and exposing corneas to the microbial suspension or by intrastromal injection. Post-inoculation, corneas were maintained for 24 and 48 h at 37 °C. After incubation, corneas were either homogenised to determine colony-forming units (CFU)/cornea or processed for histological examination using routine staining methods. Single- and mixed-species infections were compared. We observed a significant increase in CFU after 48 h compared to 24 h with S. aureus and P. aeruginosa. However, no such increase was observed in corneas infected with C. albicans or F. solani. The injection method yielded an approximately two- to 100-fold increase (p < 0.05) in the majority of organisms from infected corneas. Histology of the scalpel-wounded and injection models indicated extensive infiltration of P. aeruginosa throughout the entire cornea, with less infiltration observed for S. aureus, C. albicans and F. solani. The models also supported dual infections. Both scalpel wounding and injection methods are suitable for inducing infection of ex vivo rabbit and human cornea models. These simple and reproducible models will be useful as an alternative to in vitro and in vivo models for investigating the detection and treatment of microbial keratitis, particularly when this might be due to two infective organisms.

Cellular proliferation and regeneration following tissue damage. Progress report. [Eyes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Harding, C.V.

1976-10-01

Results are reported from a study of wound healing in tissues of the eye, particularly lens, cornea, and surrounding tissues. The reactions of these tissues to mechanical injuries, as well as injuries induced by chemotoxic agents were studied. It is postulated that a better understanding of the basic reactions of the eye to injurious agents may be of importance in the evaluation of potential environmental hazards.

[Biomaterials or Donor Tissue - What is the Future of Tissue Engenieering for Cornea Reconstruction?

PubMed

Bachmann, Björn O; Schrader, Stefan

2017-06-01

For the replacement of corneal tissue, corneal grafts or amniotic membrane are still used as a standard material. Since this is biological tissue, there is only a limited standardization regarding preparation, tissue properties and behaviour after transplantation. In addition, there is a risk of disease transmission, and the availability of both human corneas and amniotic membrane is insufficient in many regions of the world, which is why alternative biomaterials have been explored for many years now. Among the natural biomaterials, materials based on collagen or keratin provide characteristics that make them good candidates for corneal tissue replacement. However, there are still many unsolved problems, particularly regarding the degradation after implantation and the seam strength of the materials. Initial clinical studies with different biomaterials based on collagen prove their good biocompatibility to integrate and their low immunogenicity. Currently, there is no biomaterial that meets the requirements in every situation. It can be assumed that different biomaterials will be available in the future, which, depending on the underlying corneal disease, will fulfill different functions and thus make a patient- and disease-specific care possible. Georg Thieme Verlag KG Stuttgart · New York.

Epidermal growth factor in alkali-burned corneal epithelial wound healing.

PubMed

Singh, G; Foster, C S

1987-06-15

We conducted a double-masked study to evaluate the effect of epidermal growth factor on epithelial wound healing and recurrent erosions in alkali-burned rabbit corneas. Epithelial wounds 10 mm in diameter healed completely under the influence of topical epidermal growth factor, whereas the control corneas did not resurface in the center. On reversal of treatment, the previously nonhealing epithelial defects healed when treated with topical epidermal growth factor eyedrops. Conversely, the epidermal growth factor-treated and resurfaced corneas developed epithelial defects when treatment was discontinued. Histopathologic examination disclosed hyperplastic epithelium growing over the damaged stroma laden with polymorphonuclear leukocytes when treated with epidermal growth factor eyedrops, but it did not adhere to the underlying tissue. Hydropic changes were seen intracellularly as well as between the epithelial cells and the stroma.

Perilimbal sclera mechanical properties: Impact on intraocular pressure in porcine eyes

PubMed Central

Man, Xiaofei; Arroyo, Elizabeth; Dunbar, Martha; Reed, David M.; Shah, Neil; Kagemann, Larry; Kim, Wonsuk; Moroi, Sayoko E.

2018-01-01

There is extensive knowledge on the relationship of posterior scleral biomechanics and intraocular pressure (IOP) load on glaucomatous optic neuropathy; however, the role for biomechanical influence of the perilimbal scleral tissue on the aqueous humor drainage pathway, including the distal venous outflow system, and IOP regulation is not fully understood. The purpose of this work is to study the outflow characteristics of perfused porcine eyes relative to the biomechanical properties of the perilimbal sclera, the posterior sclera and the cornea. Enucleated porcine eyes from eleven different animals were perfused with surrogate aqueous at two fixed flow rates while monitoring their IOP. After perfusion, mechanical stress-strain and relaxation tests were conducted on specimens of perilimbal sclera, posterior sclera, and cornea from the same perfused eyes. Statistical analysis of the data demonstrated a strong correlation between increased tangent modulus of the perilimbal sclera tissues and increased perfusion IOP (R2 = 0.74, p = 0.0006 at lower flow rate and R2 = 0.71, p = 0.0011 at higher flow rate). In contrast, there were no significant correlations between IOP and the tangent modulus of the other tissues (Posterior sclera: R2 = 0.17 at lower flow rate and R2 = 0.30 at higher flow rate; cornea: R2 = 0.02 at lower flow rate and R2<0.01 at higher flow rate) nor the viscoelastic properties of any tissue (R2 ≤ 0.08 in all cases). Additionally, the correlation occurred for IOP and not net outflow facility (R2 ≤ 0.12 in all cases). These results provide new evidence that IOP in perfused porcine eyes is strongly influenced by the tangent modulus, sometimes called the tissue stiffness, of the most anterior portion of the sclera, i.e. the limbus. PMID:29718942

Laser Refractive Surgery

NASA Astrophysics Data System (ADS)

Wynne, James

Refractive surgery has its roots in corneal transplant surgery, first performed in 1905, where the damaged or diseased cornea of a living individual is replaced by donated corneal tissue taken from a recently deceased individual. Since the cornea has no blood supply, there is no danger of organ rejection. Recognizing the exceptional healing power of corneal tissue, ophthalmologists began to explore methods of reshaping the cornea to improve the visual acuity of patients suffering from myopia, hyperopia, and astigmatism. In 1964, a procedure known as keratomileusis was introduced. In 1974, radial keratotomy (RK) was introduced. In 1981, excimer laser surgery was discovered by the speaker and his IBM Research colleagues. In 1983, the excimer laser was used to create clean, precise incisions in the cornea of enucleated calf eyes, derived from slaughter, launching the era of laser refractive surgery, with more precise and safer techniques to correct myopia, hyperopia, and stigmatism. This talk will describe the widely practiced surgical procedures known as photorefractive keratectomy (PRK) and laser-assisted in situ keratomileusis (LASIK), which have improved the visual acuity of more than 35 million people. Most patients undergoing PRK or LASIK end up with uncorrected vision better than 20/20. In 2007, development commenced on a new procedure known as small incision lenticule extraction (SMILE), employing a femtosecond laser and no excimer laser. SMILE is promoted as minimally invasive and combining the advantages of PRK and LASIK. However, long term stability of visual acuity following SMILE surgery is yet to be determined.

Keratoconus

MedlinePlus

... condition may be due to a defect in collagen. This is the tissue that provides the shape ... the cornea so contact lenses fit better Corneal collagen cross-linking is a treatment that causes the ...

Moving epithelia: Tracking the fate of mammalian limbal epithelial stem cells.

PubMed

Di Girolamo, Nick

2015-09-01

Lineage tracing allows the destiny of a stem cell (SC) and its progeny to be followed through time. In order to track their long-term fate, SC must be permanently marked to discern their distribution, division, displacement and differentiation. This information is essential for unravelling the mysteries that govern their replenishing activity while they remain anchored within their niche microenvironment. Modern-day lineage tracing uses inducible genetic recombination to illuminate cells within embryonic, newborn and adult tissues, and the advent of powerful high-resolution microscopy has enabled the behaviour of labelled cells to be monitored in real-time in a living organism. The simple structural organization of the mammalian cornea, including its accessibility and transparency, renders it the ideal tissue to study SC fate using lineage tracing assisted by non-invasive intravital microscopy. Despite more than a century of research devoted to understanding how this tissue is maintained and repaired, many limitations and controversies continue to plague the field, including uncertainties about the specificity of current SC markers, the number of SC within the cornea, their mode of division, their location, and importantly the signals that dictate cell migration. This communication will highlight historical discoveries as well as recent developments in the corneal SC field; more specifically how the progeny of these cells are mobilised to replenish this dynamic tissue during steady-state, disease and transplantation. Also discussed is how insights gleaned from animal studies can be used to advance our knowledge of the fundamental mechanisms that govern modelling and remodelling of the human cornea in health and disease. Copyright © 2015 Elsevier Ltd. All rights reserved.

PRODUCTION OF HIGHLY-ALIGNED COLLAGEN LAMELLAE BY COMBINING SHEAR FORCE AND THIN-FILM CONFINEMENT

PubMed Central

Saeidi, Nima; Sander, Edward A.; Zareian, Ramin

2012-01-01

Load-bearing tissues owe their mechanical strength to their highly-anisotropic collagenous structure. To date, attempts to engineer mechanically strong connective tissue have failed mainly due to the lack of the ability to reproduce native collagen organization in constructs synthesized by cultured cells in vitro. The ability to influence the direction of the self-assembling collagen molecules and produce highly anisotropic structures has applications ranging from de novo engineering of complex tissues to the production of organized scaffolds for cell culture contact guidance. In this investigation we have used the simple technique of spin coating to produce highly-aligned arrays of collagen fibrils. By a simple modification of the method we have also successfully produced orthogonal collagen lamellae. Alternating collagen lamellae are frequently seen in load-bearing tissues such as cornea, annulus fibrosus, and cortical bone. Culturing of corneal fibroblasts onto aligned collagen shows that the cells adopt the organization of the fibrils. In this investigation, we observed the reversal of fibrillar growth direction or “hook” formation similar to those seen previously in a microfluidic shear-flow chamber. Although the results of this investigation clearly show that it is possible to produce small areas (O) 1 cm2 of collagen fibrils with enough alignment to guide fibroblasts, there is evidence that thin film instabilities are likely to be a significant barrier to producing organized collagen fibrils over larger areas. Successful application of this method to produce highly-controlled and organized collagenous structures will require the development of techniques to control thin film instability and will be the subject of the future work. PMID:21362500

Cromolyn Ophthalmic

MedlinePlus

... keratitis (a condition that causes swelling of the cornea [tissue in the front of the eye] that ... send a report to the Food and Drug Administration's (FDA) MedWatch Adverse Event Reporting program online (http:// ...

Corneal Wound Healing Requires IKB kinase β Signaling in Keratocytes

PubMed Central

Chen, Liang; Mongan, Maureen; Meng, Qinghang; Wang, Qin; Kao, Winston; Xia, Ying

2016-01-01

IkB kinase β (IKKβ) is a key signaling kinase for inflammatory responses, but it also plays diverse cell type-specific roles that are not yet fully understood. Here we investigated the role of IKKβ in the cornea using IkkβΔCS mice in which the Ikkβ gene was specifically deleted in the corneal stromal keratocytes. The IkkβΔCS corneas had normal morphology, transparency and thickness; however, they did not heal well from mild alkali burn injury. In contrast to the IkkβF/F corneas that restored transparency in 2 weeks after injury, over 50% of the IkkβΔCS corneas failed to fully recover. They instead developed recurrent haze with increased stromal thickness, severe inflammation and apoptosis. This pathogenesis correlated with sustained myofibroblast transformation with increased α smooth muscle actin (α-SMA) expression, higher levels of senescence β-Gal activity and scar tissue formation at the late stage of wound healing. In addition, the IkkβΔCS corneas displayed elevated expression of hemo-oxygenase-1 (HO-1), a marker of oxidative stress, and activation of stress signaling pathways with increased JNK, c-Jun and SMAD2/3 phosphorylation. These data suggest that IKKβ in keratocytes is required to repress oxidative stress and attenuate fibrogenesis and senescence in corneal wound healing. PMID:26987064

Corneal permeability for cement dust: prognosis for occupational safety

NASA Astrophysics Data System (ADS)

Kalmykov, R. V.; Popova, D. V.; Kamenskikh, T. G.; Genina, E. A.; Tuchin, V. V.; Bashkatov, A. N.

2018-02-01

The high dust content in air of a working zone causes prevalence of pathologies of the anterior segment of the eye of workers of cement production. Therefore, studying of features of cement dust impact on structure of a cornea and development of ways of eye protection from this influence is relevant. In this work experimental studies were carried out with twenty eyes of ten rabbits. OCTtomography was used to monitor the light attenuation coefficient of the cornea in vitro during the permeability of cement dust and/or keratoprotector (Systein Ultra). The permeability coefficients of the cornea for water, cement dust and keratoprotector were measured. A computer model allowing one to analyze the diffusion of these substances in the eye cornea was developed. It was shown that 1) the cement dust falling on the eye cornea caused pronounced dehydration of the tissue (thickness decreasing) and led to the increase of the attenuation coefficient, which could affect the deterioration of the eyesight of workers in the conditions of cement production; 2) the application of the keratoprotector to the eye cornea when exposed by cement dust, slowed significantly the dehydration process and did not cause the increase of the attenuation coefficient that characterized the stabilization of visual functions. At this, the keratoprotector itself did not cause dehydration and led to the decrease of the attenuation coefficient, which could allow it to be used for a long time in the order to protect the organ of vision from the negative effects of cement dust.

[Evaluation of corneal tissue by slit lamp and specular microscopy during the preservation period].

PubMed

Farias, Roberta Jansen de Mello; Kubokawa, Karen Miyuki; Schirmer, Maurício; Sousa, Luciene Barbosa de

2007-01-01

To analyze changes evaluation of corneal grafts by slit lamp and electron microscopy examination in order to ensure a donor cornea of good quality level for the patient. 203 corneas donated to the Sorocaba Eye Bank between September 2002 and February 2003 were prospectively evaluated. The evaluation was graded from 0 to 3 according to the following factors: exposure and epithelial damage, stromal opacity, Descemet folds, pleomorphism, polymegatism and guttata, endothelial snail track, edema, reflectivity. The data were correlated with time between death and preservation, endothelial cell count and donor's age. 203 corneas were evaluated. The mean age of donors was 55 years (dp= 14.8 years). The mean time between death and preservation was 9.1 h (minimum of 2 h and maximum of 25 h). Eighty-six corneas suffered worsening of evaluation, whereby 66.3% had less than 2,500 endothelial cell count and 59.3% presented time between death and preservation over 6 hours. The mean day of the grading change was the 5th. Corneal grafts from younger donors had significantly better evaluation than those of older donors. The mean time of the evaluation change was on the 5th day in 50% of the corneas, however, an expressive number of corneas suffered changes on the 6th, 7th and 8th day. Corneal graft preserved after 6 hours of death had a greater tendency to lower cell count and to decrease in evaluation grades.

Excimer laser ablation of the cornea

NASA Astrophysics Data System (ADS)

Pettit, George H.; Ediger, Marwood N.; Weiblinger, Richard P.

1995-03-01

Pulsed ultraviolet laser ablation is being extensively investigated clinically to reshape the optical surface of the eye and correct vision defects. Current knowledge of the laser/tissue interaction and the present state of the clinical evaluation are reviewed. In addition, the principal findings of internal Food and Drug Administration research are described in some detail, including a risk assessment of the laser-induced-fluorescence and measurement of the nonlinear optical properties of cornea during the intense UV irradiation. Finally, a survey is presented of the alternative laser technologies being explored for this ophthalmic application.

Nanoscale modification of porous gelatin scaffolds with chondroitin sulfate for corneal stromal tissue engineering

PubMed Central

Lai, Jui-Yang; Li, Ya-Ting; Cho, Ching-Hsien; Yu, Ting-Chun

2012-01-01

Recent studies reflect the importance of using naturally occurring biopolymers as three-dimensional corneal keratocyte scaffolds and suggest that the porous structure of gelatin materials may play an important role in controlling nutrient uptake. In the current study, the authors further consider the application of carbodiimide cross-linked porous gelatin as an alternative to collagen for corneal stromal tissue engineering. The authors developed corneal keratocyte scaffolds by nanoscale modification of porous gelatin materials with chondroitin sulfate (CS) using carbodiimide chemistry. Scanning electron microscopy/energy dispersive X-ray spectroscopy and Fourier transform infrared spectroscopy showed that the amount of covalently incorporated polysaccharide was significantly increased when the CS concentration was increased from 0% to 1.25% (w/v). In addition, as demonstrated by dimethylmethylene blue assays, the CS content in these samples was in the range of 0.078–0.149 nmol per 10 mg scaffold. When compared with their counterparts without CS treatment, various CS-modified porous gelatin membranes exhibited higher levels of water content, light transmittance, and amount of permeated nutrients but possessed lower Young’s modulus and resistance against protease digestion. The hydrophilic and mechanical properties of scaffolds modified with 0.25% CS were comparable with those of native corneas. The samples from this group were biocompatible with the rabbit corneal keratocytes and showed enhanced proliferative and biosynthetic capacity of cultured cells. In summary, the authors found that the nanoscale-level modification has influence on the characteristics and cell-material interactions of CS-containing gelatin hydrogels. Porous membranes with a CS content of 0.112 ± 0.003 nmol per 10 mg scaffold may hold potential for use in corneal stromal tissue engineering. PMID:22403490

Glaucoma Treatments and Drugs

MedlinePlus

... white part of the eye (sclera), removes a piece of trabecular meshwork (eye tissue located around the ... tiny incision in the cornea and remove a piece of the trabecular meshwork to increase fluid drainage. ...

Characterization of Ocular Iontophoretic Drug Transport of Ionic and Non-ionic Compounds in Isolated Rabbit Cornea and Conjunctiva.

PubMed

Sekijima, Hidehisa; Ehara, Junya; Hanabata, Yusuke; Suzuki, Takumi; Kimura, Soichiro; Lee, Vincent H L; Morimoto, Yasunori; Ueda, Hideo

2016-06-01

Ocular iontophoresis (IP) in isolated rabbit cornea and conjunctiva was examined in terms of transport enhancement, tissue viability and integrity using electrophysiological parameters by the Ussing-type chamber technique. Lidocaine hydrochloride (LC, a cationic compound), sodium benzoate (BA, anionic compound), and fluorescein isothiocyanate labeled dextran (molecular weight 4400 Da, FD-4, hydrophilic large compound) were used as model permeants. Direct electric current was applied at 0.5-5.0 mA/cm(2) for the cornea and 0.5-20 mA/cm(2) for the conjunctiva for 30 min. LC and BA fluxes across the cornea and conjunctiva were significantly increased by the application of electric current up to 2.3- and 2.5-fold and 4.0- and 3.4-fold, respectively, and returned to their baseline level on stopping the current. Furthermore, a much higher increase by IP application was obtained for the FD-4 transport. The increased FD-4 flux in the conjunctiva returned to baseline on stopping the current, whereas the flux in the cornea was sustained at a higher level after stopping the current. The transepithelial electric resistance of the cornea and conjunctiva was lowered by electric current application but fully recovered after stopping the current up to 2.0 mA/cm(2) for the cornea and 10 mA/cm(2) for the conjunctiva, suggesting that the corneal and conjunctival viability and integrity are maintained even after application of these current densities. These results indicate that ocular IP may be a useful non-invasive technique to achieve drug delivery of hydrophilic large molecules into the eyes.

[Structural changes in the tissues of white rats after capsaicin administration].

PubMed

Vorob'eva, N F; Kniazev, G G; Lazarev, V A; Spiridonov, V K

1997-01-01

Tissue structure of albino rat lung, skin and cornea changing after administration of capsaicin (neurotoxin isolated from red pepper) was studied using light and electron microscope. 5 mg/kg dose causes tissue swelling and microcirculatory bed reaction. 200 mg/kg dose leads to more significant dystrophic tissue alterations. Fibrosclerosis signs were found in certain cases. Microcirculatory disorders are proposed as the main reason for tissue structure alterations observed, although the mechanism of their development is still unclear.

An "All-laser" Endothelial Transplant.

PubMed

Rossi, Francesca; Canovetti, Annalisa; Malandrini, Alex; Lenzetti, Ivo; Pini, Roberto; Menabuoni, Luca

2015-07-06

The "all laser" assisted endothelial keratoplasty is a procedure that is performed with a femtosecond laser used to cut the donor tissue at an intended depth, and a near infrared diode laser to weld the corneal tissue. The proposed technique enables to reach the three main goals in endothelial keratoplasty: a precise control in the thickness of the donor tissue; its easy insertion in the recipient bed and a reduced risk of donor lenticule dislocation. The donor cornea thickness is measured in the surgery room with optical coherence tomography (OCT), in order to correctly design the donor tissue dimensions. A femtosecond laser is used to cut the donor cornea. The recipient eye is prepared by manual stripping of the descemetic membrane. The donor endothelium is inserted into a Busin-injector, the peripheral inner side is stained with a proper chromophore (a water solution of Indocyanine Green) and then it is pulled in the anterior chamber. The transplanted tissue is placed in the final and correct location and then diode laser welding is induced from outside the eyeball. The procedure has been performed on more than 15 patients evidencing an improvement in surgery performances, with a good recovery of visual acuity and a reduced donor lenticule dislocation event.

Diffusion and Monod kinetics model to determine in vivo human corneal oxygen-consumption rate during soft contact lens wear

PubMed Central

Del Castillo, Luis F.; da Silva, Ana R. Ferreira; Hernández, Saul I.; Aguilella, M.; Andrio, Andreu; Mollá, Sergio; Compañ, Vicente

2014-01-01

Purpose We present an analysis of the corneal oxygen consumption Qc from non-linear models, using data of oxygen partial pressure or tension (pO2) obtained from in vivo estimation previously reported by other authors.1 Methods Assuming that the cornea is a single homogeneous layer, the oxygen permeability through the cornea will be the same regardless of the type of lens that is available on it. The obtention of the real value of the maximum oxygen consumption rate Qc,max is very important because this parameter is directly related with the gradient pressure profile into the cornea and moreover, the real corneal oxygen consumption is influenced by both anterior and posterior oxygen fluxes. Results Our calculations give different values for the maximum oxygen consumption rate Qc,max, when different oxygen pressure values (high and low pO2) are considered at the interface cornea-tears film. Conclusion Present results are relevant for the calculation on the partial pressure of oxygen, available at different depths into the corneal tissue behind contact lenses of different oxygen transmissibility. PMID:25649636

Novel micelle carriers for cyclosporin A topical ocular delivery: in vivo cornea penetration, ocular distribution and efficacy studies.

PubMed

Di Tommaso, Claudia; Bourges, Jean-Louis; Valamanesh, Fatemeh; Trubitsyn, Gregory; Torriglia, Alicia; Jeanny, Jean-Claude; Behar-Cohen, Francine; Gurny, Robert; Möller, Michael

2012-06-01

Cornea transplantation is one of the most performed graft procedures worldwide with an impressive success rate of 90%. However, for "high-risk" patients with particular ocular diseases in addition to the required surgery, the success rate is drastically reduced to 50%. In these cases, cyclosporin A (CsA) is frequently used to prevent the cornea rejection by a systemic treatment with possible systemic side effects for the patients. To overcome these problems, it is a challenge to prepare well-tolerated topical CsA formulations. Normally high amounts of oils or surfactants are needed for the solubilization of the very hydrophobic CsA. Furthermore, it is in general difficult to obtain ocular therapeutic drug levels with topical instillations due to the corneal barriers that efficiently protect the intraocular structures from foreign substances thus also from drugs. The aim of this study was to investigate in vivo the effects of a novel CsA topical aqueous formulation. This formulation was based on nanosized polymeric micelles as drug carriers. An established rat model for the prevention of cornea graft rejection after a keratoplasty procedure was used. After instillation of the novel formulation with fluorescent labeled micelles, confocal analysis of flat-mounted corneas clearly showed that the nanosized carriers were able to penetrate into all corneal layers. The efficacy of a 0.5% CsA micelle formulation was tested and compared to a physiological saline solution and to a systemic administration of CsA. In our studies, the topical CsA treatment was carried out for 14 days, and the three parameters (a) cornea transparency, (b) edema, and (c) neovascularization were evaluated by clinical observation and scoring. Compared to the control group, the treated group showed a significant higher cornea transparency and significant lower edema after 7 and 13 days of the surgery. At the end point of the study, the neovascularization was reduced by 50% in the CsA-micelle treated animals. The success rate of cornea graft transplantation was 73% in treated animals against 25% for the control group. This result was as good as observed for a systemic CsA treatment in the same animal model. This new formulation has the same efficacy like a systemic treatment but without the serious CsA systemic side effects. Ocular drug levels of transplanted and healthy rat eyes were dosed by UPLC/MS and showed a high CsA value in the cornea (11710 ± 7530 ng(CsA)/g(tissue) and 6470 ± 1730 ng(CsA)/g(tissue), respectively). In conclusion, the applied formulation has the capacity to overcome the ocular surface barriers, the micelles formed a drug reservoir in the cornea from, where a sustained release of CsA can take place. This novel formulation for topical application of CsA is clearly an effective and well-tolerated alternative to the systemic treatment for the prevention of corneal graft rejection. Copyright © 2012 Elsevier B.V. All rights reserved.

Assessment of the Accuracy and Cut-Failure Rates of Eye Bank-Cut Corneas for Use in Endothelial Keratoplasty: A Comparison of Outcomes Between 2010 and 2013.

PubMed

Katzman, Lee R; Hoover, Caroline K; Khalifa, Yousuf M; Jeng, Bennie H

2015-11-01

To evaluate the accuracy of eye bank-prepared precut donor corneas over time by comparing cut-failure rates and corneal thickness measurements in 2010 and 2013. A total of 2511 human corneas cut by a technician-operated mechanical microkeratome intended for endothelial keratoplasty were evaluated prospectively at one large eye bank facility in 2010 and in 2013. The endothelium was evaluated by slit lamp, and specular microscopy both before and after cutting was performed. Graft thickness as measured by pachymetry and/or optical coherence tomography was collected to assess the accuracy of the cut tissue. Cut-failure rates were compared between normal donor tissue and tissue with significant preexisting scarring. The combined cut-failure rate in 2010 and 2013 was 2.3% (23/1000) and 1.6% (24/1511), respectively (P = 0.23). The cut-failure rate among normal tissue in 2010 and 2013 was 2.0% (19/927) and 1.4% (19/1400), respectively (P = 0.24). The cut-failure rate among previously scarred tissue in 2010 and 2013 was 5.5% (4/73) and 4.5% (5/111), respectively (P = 0.74). The mean surgeon-requested graft thickness was 144.7 μm (range 100-150, SD 13.6) and 127.2 μm (range 75-150, SD 25.2) in 2010 and 2013, respectively (P < 0.0001). The mean deviation from target graft thickness was 21.3 μm (SD 16.3) and 13.6 μm (SD 12.5) in 2010 and 2013, respectively (P < 0.0001). From 2010 to 2013, the combined cut-failure rates trended toward improvement, while the accuracy of graft thickness improved. This study suggests that the accuracy and success rates of tissue preparation for endothelial keratoplasty improve with experience and volume.

Computer simulation of photorefractive keratectomy for the correction of myopia and hyperopia

NASA Astrophysics Data System (ADS)

Pinault, Pascal; 'Huillier, J. P.

1996-01-01

Photorefractive keratectomy (PRK) performed by means of the 193 nm excimer laser has stimulated considerable interest in the ophthalmic community because this new procedure has the potential to correct myopia, hyperopia, and astigmatism. The use of a laser beam to remove a controlled amount of tissue from the cornea implies that both the energy density of the laser beam and the target removed rate are accurately known. In addition, the required tissue ablation profile to achieve refractive correction must be predicted by optical calculations. This paper investigates: (1) Optical computations based on raytracing model to determine what anterior profile of cornea is needed postoperatively for ametropia. (2) Maximal depth of the removed corneal tissue against the ablation zone treated. And (3) the thickness of ablated corneal lenticule at any distance from the optical axis. Relationships between these data are well fitted by polynomial regressive curves in order to be useful as an algorithm in the computer-controlled delivery of the ArF laser beam.

High-intensity corneal collagen crosslinking with riboflavin and UVA in rat cornea.

PubMed

Zhu, Yirui; Reinach, Peter S; Zhu, Hanlei; Tan, Qiufan; Zheng, Qinxiang; Qu, Jia; Chen, Wei

2017-01-01

Corneal collagen cross-linking (CXL) halts human corneal ectasias progression by increasing stromal mechanical stiffness. Although some reports describe that this procedure is effective in dealing with some infectious and immunologic corneal thinning diseases, there is a need for more animal models whose corneal thickness more closely resemble those occurring in these patients. To meet this need, we describe here high-intensity protocols that are safe and effective for obtaining CXL in rat corneas. Initially, a range of potentially effective UVA doses were evaluated based on their effectiveness in increasing tissue enzymatic resistance to dissolution. At UVA doses higher than a threshold level of 0.54 J/cm2, resistance to enzymatic digestion increased relative to that in non-irradiated corneas. Based on the theoretical threshold CXL dose, a CXL regimen was established in which the UVA tissue irradiance was 9 mW/cm2, which was delivered at doses of either 2.16, 2.7 or 3.24 J/cm2. Their dose dependent effects were evaluated on ocular surface morphological integrity, keratocyte apoptotic frequency, tissue thickness and endothelial cell layer density. Doses of 2.16 and 2.7 J/cm2 transiently decreased normal corneal transparency and increased thickness. These effects were fully reversed after 14 days. In contrast, 3.24 J/cm2 had more irreversible side effects. Three days after treatment, apoptotic frequency in the CXL-2.16 group was lower than that at higher doses. Endothelial cell losses remained evident only in the CXL-3.24 group at 42 days posttreatment. Stromal fiber thickening was evident in all the CXL-treated groups. We determined both the threshold UVA dose using the high-intensity CXL procedure and identified an effective dose range that provides optimal CXL with minimal transient side effects in the rat cornea. These results may help to provide insight into how to improve the CXL outcome in patients afflicted with a severe corneal thinning disease.

Glutathione-related enzymes and the eye.

PubMed

Ganea, Elena; Harding, John J

2006-01-01

Glutathione and the related enzymes belong to the defence system protecting the eye against chemical and oxidative stress. This review focuses on GSH and two key enzymes, glutathione reductase and glucose-6-phosphate dehydrogenase in lens, cornea, and retina. Lens contains a high concentration of reduced glutathione, which maintains the thiol groups in the reduced form. These contribute to lens complete transparency as well as to the transparent and refractive properties of the mammalian cornea, which are essential for proper image formation on the retina. In cornea, gluthatione also plays an important role in maintaining normal hydration level, and in protecting cellular membrane integrity. In retina, glutathione is distributed in the different types of retinal cells. Intracellular enzyme, glutathione reductase, involved in reducing the oxidized glutathione has been found at highest activity in human and primate lenses, as compared to other species. Besides the enzymes directly involved in maintaining the normal redox status of the cell, glucose-6-phosphate dehydrogenase which catalyzes the first reaction of the pentose phosphate pathway, plays a key role in protection of the eye against reactive oxygen species. Cornea has a high activity of the pentose phosphate pathway and glucose-6-phosphate dehydrogenase activity. Glycation, the non-enzymic reaction between a free amino group in proteins and a reducing sugar, slowly inactivates gluthathione-related and other enzymes. In addition, glutathione can be also glycated. The presence of glutathione, and of the related enzymes has been also reported in other parts of the eye, such as ciliary body and trabecular meshwork, suggesting that the same enzyme systems are present in all tissues of the eye to generate NADPH and to maintain gluthatione in the reduced form. Changes of glutathione and related enzymes activity in lens, cornea, retina and other eye tissues, occur with ageing, cataract, diabetes, irradiation and administration of some drugs.

The impact of intraocular pressure on elastic wave velocity estimates in the crystalline lens.

PubMed

Park, Suhyun; Yoon, Heechul; Larin, Kirill V; Emelianov, Stanislav Y; Aglyamov, Salavat R

2016-12-20

Intraocular pressure (IOP) is believed to influence the mechanical properties of ocular tissues including cornea and sclera. The elastic properties of the crystalline lens have been mainly investigated with regard to presbyopia, the age-related loss of accommodation power of the eye. However, the relationship between the elastic properties of the lens and IOP remains to be established. The objective of this study is to measure the elastic wave velocity, which represents the mechanical properties of tissue, in the crystalline lens ex vivo in response to changes in IOP. The elastic wave velocities in the cornea and lens from seven enucleated bovine globe samples were estimated using ultrasound shear wave elasticity imaging. To generate and then image the elastic wave propagation, an ultrasound imaging system was used to transmit a 600 µs pushing pulse at 4.5 MHz center frequency and to acquire ultrasound tracking frames at 6 kHz frame rate. The pushing beams were separately applied to the cornea and lens. IOP in the eyeballs was varied from 5 to 50 mmHg. The results indicate that while the elastic wave velocity in the cornea increased from 0.96  ±  0.30 m s -1 to 6.27  ±  0.75 m s -1 as IOP was elevated from 5 to 50 mmHg, there were insignificant changes in the elastic wave velocity in the crystalline lens with the minimum and the maximum speeds of 1.44  ±  0.27 m s -1 and 2.03  ±  0.46 m s -1 , respectively. This study shows that ultrasound shear wave elasticity imaging can be used to assess the biomechanical properties of the crystalline lens noninvasively. Also, it was observed that the dependency of the crystalline lens stiffness on the IOP was significantly lower in comparison with that of cornea.

The effect of riboflavin/UVA collagen cross-linking therapy on the structure and hydrodynamic behaviour of the ungulate and rabbit corneal stroma.

PubMed

Hayes, Sally; Kamma-Lorger, Christina S; Boote, Craig; Young, Robert D; Quantock, Andrew J; Rost, Anika; Khatib, Yasmeen; Harris, Jonathan; Yagi, Naoto; Terrill, Nicholas; Meek, Keith M

2013-01-01

To examine the effect of riboflavin/UVA corneal crosslinking on stromal ultrastructure and hydrodynamic behaviour. One hundred and seventeen enucleated ungulate eyes (112 pig and 5 sheep) and 3 pairs of rabbit eyes, with corneal epithelium removed, were divided into four treatment groups: Group 1 (28 pig, 2 sheep and 3 rabbits) were untreated; Group 2 (24 pig) were exposed to UVA light (3.04 mW/cm(2)) for 30 minutes and Group 3 (29 pig) and Group 4 (31 pig, 3 sheep and 3 rabbits) had riboflavin eye drops applied to the corneal surface every 5 minutes for 35 minutes. Five minutes after the initial riboflavin instillation, the corneas in Group 4 experienced a 30 minute exposure to UVA light (3.04 mW/cm(2)). X-ray scattering was used to obtain measurements of collagen interfibrillar spacing, spatial order, fibril diameter, D-periodicity and intermolecular spacing throughout the whole tissue thickness and as a function of tissue depth in the treated and untreated corneas. The effect of each treatment on the hydrodynamic behaviour of the cornea (its ability to swell in saline solution) and its resistance to enzymatic digestion were assessed using in vitro laboratory techniques. Corneal thickness decreased significantly following riboflavin application (p<0.01) and also to a lesser extent after UVA exposure (p<0.05). With the exception of the spatial order factor, which was higher in Group 4 than Group 1 (p<0.01), all other measured collagen parameters were unaltered by cross-linking, even within the most anterior 300 microns of the cornea. The cross-linking treatment had no effect on the hydrodynamic behaviour of the cornea but did cause a significant increase in its resistance to enzymatic digestion. It seems likely that cross-links formed during riboflavin/UVA therapy occur predominantly at the collagen fibril surface and in the protein network surrounding the collagen.

The Effect of Riboflavin/UVA Collagen Cross-linking Therapy on the Structure and Hydrodynamic Behaviour of the Ungulate and Rabbit Corneal Stroma

PubMed Central

Hayes, Sally; Kamma-Lorger, Christina S.; Boote, Craig; Young, Robert D.; Quantock, Andrew J.; Rost, Anika; Khatib, Yasmeen; Harris, Jonathan; Yagi, Naoto; Terrill, Nicholas; Meek, Keith M.

2013-01-01

Purpose To examine the effect of riboflavin/UVA corneal crosslinking on stromal ultrastructure and hydrodynamic behaviour. Methods One hundred and seventeen enucleated ungulate eyes (112 pig and 5 sheep) and 3 pairs of rabbit eyes, with corneal epithelium removed, were divided into four treatment groups: Group 1 (28 pig, 2 sheep and 3 rabbits) were untreated; Group 2 (24 pig) were exposed to UVA light (3.04 mW/cm2) for 30 minutes and Group 3 (29 pig) and Group 4 (31 pig, 3 sheep and 3 rabbits) had riboflavin eye drops applied to the corneal surface every 5 minutes for 35 minutes. Five minutes after the initial riboflavin instillation, the corneas in Group 4 experienced a 30 minute exposure to UVA light (3.04 mW/cm2). X-ray scattering was used to obtain measurements of collagen interfibrillar spacing, spatial order, fibril diameter, D-periodicity and intermolecular spacing throughout the whole tissue thickness and as a function of tissue depth in the treated and untreated corneas. The effect of each treatment on the hydrodynamic behaviour of the cornea (its ability to swell in saline solution) and its resistance to enzymatic digestion were assessed using in vitro laboratory techniques. Results Corneal thickness decreased significantly following riboflavin application (p<0.01) and also to a lesser extent after UVA exposure (p<0.05). With the exception of the spatial order factor, which was higher in Group 4 than Group 1 (p<0.01), all other measured collagen parameters were unaltered by cross-linking, even within the most anterior 300 microns of the cornea. The cross-linking treatment had no effect on the hydrodynamic behaviour of the cornea but did cause a significant increase in its resistance to enzymatic digestion. Conclusions It seems likely that cross-links formed during riboflavin/UVA therapy occur predominantly at the collagen fibril surface and in the protein network surrounding the collagen. PMID:23349690

Fibroblast growth factor receptor 2 (FGFR2) is required for corneal epithelial cell proliferation and differentiation during embryonic development.

PubMed

Zhang, Jinglin; Upadhya, Dinesh; Lu, Lin; Reneker, Lixing W

2015-01-01

Fibroblast growth factors (FGFs) play important roles in many aspects of embryonic development. During eye development, the lens and corneal epithelium are derived from the same surface ectodermal tissue. FGF receptor (FGFR)-signaling is essential for lens cell differentiation and survival, but its role in corneal development has not been fully investigated. In this study, we examined the corneal defects in Fgfr2 conditional knockout mice in which Cre expression is activated at lens induction stage by Pax6 P0 promoter. The cornea in LeCre, Fgfr2(loxP/loxP) mice (referred as Fgfr2(CKO)) was analyzed to assess changes in cell proliferation, differentiation and survival. We found that Fgfr2(CKO) cornea was much thinner in epithelial and stromal layer when compared to WT cornea. At embryonic day 12.5-13.5 (E12.5-13.5) shortly after the lens vesicle detaches from the overlying surface ectoderm, cell proliferation (judged by labeling indices of Ki-67, BrdU and phospho-histone H3) was significantly reduced in corneal epithelium in Fgfr2(CKO) mice. At later stage, cell differentiation markers for corneal epithelium and underlying stromal mesenchyme, keratin-12 and keratocan respectively, were not expressed in Fgfr2(CKO) cornea. Furthermore, Pax6, a transcription factor essential for eye development, was not present in the Fgfr2(CKO) mutant corneal epithelial at E16.5 but was expressed normally at E12.5, suggesting that FGFR2-signaling is required for maintaining Pax6 expression in this tissue. Interestingly, the role of FGFR2 in corneal epithelial development is independent of ERK1/2-signaling. In contrast to the lens, FGFR2 is not required for cell survival in cornea. This study demonstrates for the first time that FGFR2 plays an essential role in controlling cell proliferation and differentiation, and maintaining Pax6 levels in corneal epithelium via ERK-independent pathways during embryonic development.

Potential for Hospital Based Corneal Retreival in Hassan District Hospital

PubMed Central

Melsakkare, Suresh Ramappa; Manipur, Sahana R.; Acharya, Pavana; Ramamurthy, Lakshmi Bomalapura

2015-01-01

Context In developing countries, corneal diseases are the second leading cause of blindness. This corneal blindness can be treated through corneal transplantation. Though the present infrastructure is strong enough to increase keratoplasty numbers at a required rate, India has largest corneal blind population in the world. So a constant supply of high quality donor corneal tissue is the key factor for reduction of prevalence of corneal blindness. Considering the magnitude of corneal blindness and shortage of donor cornea, there is a huge gap in the demand and supply. Aim To study the potential for hospital based retrieval of donor corneal tissue in Hassan district hospital after analysing the indicated and contraindicated causes of deaths, so that hospital corneal retrieval program in Hassan district hospital can be planned. Materials and Methods The cross-sectional, retrospective and record-based study included all hospital deaths with age group more than two years occurred during one year period (January 2014 to December 2014). Data regarding demographic profile, cause of death, treatment given and presence of any systemic diseases were collected. The causes of deaths which are contraindicated for the retrieval of corneas were analysed and noted. The contraindications were based on the NPCB guidelines for standard of eye banking in India 2009. Results Out of 855 deaths, number of deaths in males (565) was greater than females (290). Numbers of deaths were highest between 41-60 years age group (343). Deaths due to HIV, septicaemia, meningitis, encephalitis, disseminated malignancies were contraindicated for corneal retrieval. Corneas could be retrieved from 736 deaths out of 855. Potential for corneal retrieval in a period of one year in Hassan District hospital was 86%. Conclusion Hospital corneal retrieval program has got a great potential to bridge the gap between the need for the cornea and actually collected corneas which will contribute enormously in eliminating corneal blindness. In present study there was 86% potential for corneal retrieval among the hospital deaths. PMID:26435976

Vision Restoration with a Collagen Cross-linked Boston Keratoprosthesis Unit

DTIC Science & Technology

2017-09-01

Tissue Bank International, the tissue bank used for this study changed their company name to KeraLink International. The name was changed to better...solution to KeraLink International, and Dr. Joseph Ciolino traveled to the tissue bank to train staff on the Cross-Linking procedure. KeraLink is now... tissue by collagen-crosslinking the cornea graft ex vivo using vitamin B2 (riboflavin) and ultraviolet light. The overall objective of this study is

Measurement and design of refractive corrections using ultrafast laser-induced intra-tissue refractive index shaping in live cats

NASA Astrophysics Data System (ADS)

Brooks, Daniel R.; Wozniak, Kaitlin T.; Knox, Wayne; Ellis, Jonathan D.; Huxlin, Krystel R.

2018-02-01

Intra-Tissue Refractive Index Shaping (IRIS) uses a 405 nm femtosecond laser focused into the stromal region of the cornea to induce a local refractive index change through multiphoton absorption. This refractive index change can be tailored through scanning of the focal region and variations in laser power to create refractive structures, such as gradient index lenses for visual refractive correction. Previously, IRIS was used to create 2.5 mm wide, square, -1 D cylindrical refractive structures in living cats. In the present work, we first wrote 400 μm wide bars of refractive index change at varying powers in enucleated cat globes using a custom flexure-based scanning system. The cornea and surrounding sclera were then removed and mounted into a wet cell. The induced optical phase change was measured with a Mach- Zehnder Interferometer (MZI), and appeared as fringe displacement, whose magnitude was proportional to the refractive index change. The interferograms produced by the MZI were analyzed with a Fourier Transform based algorithm in order to extract the phase change. This provided a phase change versus laser power calibration, which was then used to design the scanning and laser power distribution required to create -1.5 D cylindrical Fresnel lenses in cat cornea covering an area 6 mm in diameter. This prescription was inscribed into the corneas of one eye each of two living cats, under surgical anesthesia. It was then verified in vivo by contrasting wavefront aberration measurements collected pre- IRIS with those obtained over six months post-IRIS using a Shack-Hartmann wavefront sensor.

Sequence, molecular properties, and chromosomal mapping of mouse lumican

NASA Technical Reports Server (NTRS)

Funderburgh, J. L.; Funderburgh, M. L.; Hevelone, N. D.; Stech, M. E.; Justice, M. J.; Liu, C. Y.; Kao, W. W.; Conrad, G. W.; Spooner, B. S. (Principal Investigator)

1995-01-01

PURPOSE. Lumican is a major proteoglycan of vertebrate cornea. This study characterizes mouse lumican, its molecular form, cDNA sequence, and chromosomal localization. METHODS. Lumican sequence was determined from cDNA clones selected from a mouse corneal cDNA expression library using a bovine lumican cDNA probe. Tissue expression and size of lumican mRNA were determined using Northern hybridization. Glycosidase digestion followed by Western blot analysis provided characterization of molecular properties of purified mouse corneal lumican. Chromosomal mapping of the lumican gene (Lcn) used Southern hybridization of a panel of genomic DNAs from an interspecific murine backcross. RESULTS. Mouse lumican is a 338-amino acid protein with high-sequence identity to bovine and chicken lumican proteins. The N-terminus of the lumican protein contains consensus sequences for tyrosine sulfation. A 1.9-kb lumican mRNA is present in cornea and several other tissues. Antibody against bovine lumican reacted with recombinant mouse lumican expressed in Escherichia coli and also detected high molecular weight proteoglycans in extracts of mouse cornea. Keratanase digestion of corneal proteoglycans released lumican protein, demonstrating the presence of sulfated keratan sulfate chains on mouse corneal lumican in vivo. The lumican gene (Lcn) was mapped to the distal region of mouse chromosome 10. The Lcn map site is in the region of a previously identified developmental mutant, eye blebs, affecting corneal morphology. CONCLUSIONS. This study demonstrates sulfated keratan sulfate proteoglycan in mouse cornea and describes the tools (antibodies and cDNA) necessary to investigate the functional role of this important corneal molecule using naturally occurring and induced mutants of the murine lumican gene.

Genetics Home Reference: Fuchs endothelial dystrophy

MedlinePlus

... a protein that is part of type VIII collagen. Type VIII collagen is largely found within the cornea, surrounding the endothelial cells. Specifically, type VIII collagen is a major component of a tissue at ...

Corneal and skin laser exposures from 1540-nm laser pulses

NASA Astrophysics Data System (ADS)

Johnson, Thomas E.; Mitchell, Michael A.; Rico, Pedro J.; Fletcher, David J.; Eurell, Thomas E.; Roach, William P.

2000-06-01

Mechanisms of tissue damage are investigated for skin and cornea exposures from 1540 nm ('eye safe') laser single pulses of 0.8 milli-seconds. New skin model data point out the advantages of using the Yucatan mini-pig versus the Yorkshire pig for in-vivo skin laser exposures. Major advantages found include similarities in thickness and melanin content when compared with human skin. Histology from Yucatan mini-pig skin exposures and the calculation of an initial ED50 threshold indicate that the main photon tissue interaction may not be solely due to water absorption. In-vitro corneal equivalents compared well with in-vivo rabbit cornea exposure under similar laser conditions. In-vivo and in-vitro histology show that initial energy deposition leading to damage occurs intrastromally, while epithelial cells show no direct injury due to laser light absorption.

Limited Versus Total Epithelial Debridement Ocular Surface Injury: Live Fluorescence Imaging of Hemangiogenesis and Lymphangiogenesis in Prox1-GFP/Flk1::myr-mCherry Mice

PubMed Central

Chang, Jin-Hong; Putra, Ilham; Huang, Yu-hui; Chang, Michael; Han, Kyuyeon; Zhong, Wei; Gao, Xinbo; Wang, Shuangyong; Dugas-Ford, Jennifer; Nguyen, Tara; Hong, Young-Kwon; Azar, Dimitri T.

2016-01-01

Background Immunohistochemical staining experiments have shown that both hemangiogenesis and lymphangiogenesis occur following severe corneal and conjunctival injury and that the neovascularization of the cornea often has severe visual consequences. To better understand how hemangiogenesis and lymphangiogenesis are induced by different degrees of ocular injury, we investigated patterns of injury-induced corneal neovascularization in live Prox1-GFP/Flk1::myr-mCherry mice, in which blood and lymphatic vessels can be imaged simultaneously in vivo. Methods The eyes of Prox1-GFP/Flk1::myr-mCherry mice were injured according to four models based on epithelial debridement of the: A) central cornea (a 1.5-mm-diameter circle of tissue over the corneal apex), B) total cornea, C) bulbar conjunctiva, and D) cornea+bulbar conjunctiva. Corneal blood and lymphatic vessels were imaged on days 0, 3, 7, and 10 post-injury, and the percentages of the cornea containing blood and lymphatic vessels were calculated. Results Neither central corneal nor bulbar conjunctival debridement resulted in significant vessel growth in the mouse cornea, whereas total corneal and corneal+bulbar conjunctival debridement did. On day 10 in the central cornea, total cornea, bulbar conjunctiva, and corneal+bulbar conjunctival epithelial debridement models, the percentage of the corneal surface that was occupied by blood vessels (hemangiogenesis) was 1.9±0.8%, 7.14±2.4%, 2.29±1%, and 15.05±2.14%, respectively, and the percentage of the corneal surface that was occupied by lymphatic vessels (lymphangiogenesis) was 2.45±1.51%, 4.85±0.95%, 2.95±1.27%, and 4.15±3.85%, respectively. Conclusions Substantial corneal debridement was required to induce corneal neovascularization in the mouse cornea, and the corneal epithelium may therefore be partially responsible for maintaining corneal avascularity. General significance Our study demonstrates that GFP/Flk1::myr-mCherry mice are a useful model for studying coordinated hemangiogenic and lymphangiogenic responses. PMID:27233452

Corneal donor tissue preparation for endothelial keratoplasty.

PubMed

Woodward, Maria A; Titus, Michael; Mavin, Kyle; Shtein, Roni M

2012-06-12

Over the past ten years, corneal transplantation surgical techniques have undergone revolutionary changes. Since its inception, traditional full thickness corneal transplantation has been the treatment to restore sight in those limited by corneal disease. Some disadvantages to this approach include a high degree of post-operative astigmatism, lack of predictable refractive outcome, and disturbance to the ocular surface. The development of Descemet's stripping endothelial keratoplasty (DSEK), transplanting only the posterior corneal stroma, Descemet's membrane, and endothelium, has dramatically changed treatment of corneal endothelial disease. DSEK is performed through a smaller incision; this technique avoids 'open sky' surgery with its risk of hemorrhage or expulsion, decreases the incidence of postoperative wound dehiscence, reduces unpredictable refractive outcomes, and may decrease the rate of transplant rejection. Initially, cornea donor posterior lamellar dissection for DSEK was performed manually resulting in variable graft thickness and damage to the delicate corneal endothelial tissue during tissue processing. Automated lamellar dissection (Descemet's stripping automated endothelial keratoplasty, DSAEK) was developed to address these issues. Automated dissection utilizes the same technology as LASIK corneal flap creation with a mechanical microkeratome blade that helps to create uniform and thin tissue grafts for DSAEK surgery with minimal corneal endothelial cell loss in tissue processing. Eye banks have been providing full thickness corneas for surgical transplantation for many years. In 2006, eye banks began to develop methodologies for supplying precut corneal tissue for endothelial keratoplasty. With the input of corneal surgeons, eye banks have developed thorough protocols to safely and effectively prepare posterior lamellar tissue for DSAEK surgery. This can be performed preoperatively at the eye bank. Research shows no significant difference in terms of the quality of the tissue or patient outcomes using eye bank precut tissue versus surgeon-prepared tissue for DSAEK surgery. For most corneal surgeons, the availability of precut DSAEK corneal tissue saves time and money, and reduces the stress of performing the donor corneal dissection in the operating room. In part because of the ability of the eye banks to provide high quality posterior lamellar corneal in a timely manner, DSAEK has become the standard of care for surgical management of corneal endothelial disease. The procedure that we are describing is the preparation of the posterior lamellar cornea at the eye bank for transplantation in DSAEK surgery (Figure 1).

In vivo three-dimensional confocal laser scanning microscopy of the epithelial nerve structure in the human cornea.

PubMed

Stachs, Oliver; Zhivov, Andrey; Kraak, Robert; Stave, Joachim; Guthoff, Rudolf

2007-04-01

Evaluation of a new method for in vivo visualization of the distribution and morphology of human anterior corneal nerves. The anterior cornea was examined to a depth of 100 microm in four human volunteers with a confocal laser scanning microscope (CLSM) using a Rostock Cornea Module (developed in house) attached to a Heidelberg Retina Tomograph II (Heidelberg Engineering, Germany). Optical sections were digitally reconstructed in 3D using AMIRA (TGS Inc., USA). The scanned volumes had a greatest size of 300 x 300 x 40 microm and voxel size of 0.78 x 0.78 x 0.95 microm. The spatial arrangement of the epithelium, nerves and keratocytes was visualized by in vivo 3D-CLSM. The 3D-reconstruction of the volunteers' corneas in combination with the oblique sections gave a picture of the nerves in the central human cornea. Thin nerves run in the subepithelial plexus aligned parallel to Bowman's layer and are partially interconnected. The diameter of these fibres varied between 1.0 and 5 microm. Thick fibres rose out of the deeper stroma. The diameter of the main nerve trunks was 12+/-2 microm. Branches penetrating the anterior epithelial cell layer could not be visualized. 3D-CLSM allows analysis of the spatial arrangement of the anterior corneal nerves and visualization of the epithelium and keratocytes in the living human cornea. The developed method provides a basis for further studies of alterations of the cellular arrangement and epithelial innervation in corneal disease. This may help to clarify alterations of nerve fibre patterns under various clinical and experimental conditions.

MUC19 expression in human ocular surface and lacrimal gland and its alteration in Sjögren syndrome patients.

PubMed

Yu, D F; Chen, Y; Han, J M; Zhang, H; Chen, X P; Zou, W J; Liang, L Y; Xu, C C; Liu, Z G

2008-02-01

This study investigated the expression of MUC19, a newly discovered gel-forming mucin gene, in normal human lacrimal functional unit components and its alteration in Sjögren syndrome patients. Real-time PCR and immunohistochemistry were performed to determine the expression of MUC19 and MUC5AC in human cornea, conjunctiva, and lacrimal gland tissues. Conjunctival impression cytology specimens were collected from normal control subjects and Sjögren syndrome patients for Real-time PCR, PAS staining, and immunohistochemistry assays. In addition, conjunctiva biopsy specimens from both groups were examined for the expression differences of MUC19 and MUC5AC at both mRNA and protein level. The MUC19 mRNA was found to be present in cornea, conjunctiva and lacrimal gland tissues. The immunohistochemical staining of mucins showed that MUC19 was expressed in epithelial cells from corneal, conjunctival, and lacrimal gland tissues. In contrast, MUC5AC mRNA was only present in conjunctiva and lacrimal gland tissues, but not in cornea. Immunostaining demonstrates the co-staining of MUC19 and MUC5AC in conjunctival goblet cells. Consistent with the significant decrease of mucous secretion, both MUC19 and MUC5AC were decreased in conjunctiva of Sjögren syndrome patients compared to normal subjects. Considering the contribution of gel-forming mucins to the homeostasis of the ocular surface, the decreased expression of MUC19 and MUC5AC in Sjögren syndrome patients suggested that these mucins may be involved in the disruption of the ocular surface homeostasis in this disease.

Disposition and metabolism of a novel prostanoid antiglaucoma medication, tafluprost, following ocular administration to rats.

PubMed

Fukano, Y; Kawazu, K

2009-08-01

The disposition and metabolism of tafluprost, an ester prodrug of the 15,15-difluoro-prostaglandin F(2alpha) antiglaucoma agent, have been studied in rats after ocular administration. Radioactivity was absorbed very rapidly into the eye and systemic circulation after a single ocular dose of 0.005% [(3)H]tafluprost ophthalmic solution, with maximum levels in plasma and most eye tissues occurring within 15 min. The absorption ratio of radioactivity was approximately 75%, suggesting the high availability of ocular administration of tafluprost. Approximately 10% of the dose was present in cornea at this time, and radioactivity concentrations in this tissue exceeded those in aqueous humor and iris/ciliary body throughout the 24-h study period. After repeated daily ocular doses, radioactivity levels remained greatest in cornea, followed by iris/ciliary body that replaced aqueous humor as the eye tissue containing the second highest radioactivity concentration. In female rats, radioactivity was excreted equally between urine and feces after a single ocular dose, whereas in male rats more was excreted in feces, reflecting the greater biliary excretion in males rats (50% dose) compared with females rats (33% dose). Tafluprost was extensively metabolized in the rat, such that intact prodrug was not detected in plasma, tissues, or excreta by radio-high-performance liquid chromatography. On the other hand, the active moiety, tafluprost acid, was the only noteworthy radioactive component in cornea, aqueous humor, and iris/ciliary body for at least 8 h after the ocular dose, and it was also a major plasma metabolite in early time points. The gender differences in conjugation reactions resulted in the differences in the excretion.

Dynamic OCT measurement of corneal deformation by an air puff in normal and cross-linked corneas

PubMed Central

Dorronsoro, Carlos; Pascual, Daniel; Pérez-Merino, Pablo; Kling, Sabine; Marcos, Susana

2012-01-01

A new technique is presented for the non-invasive imaging of the dynamic response of the cornea to an air puff inducing a deformation. A spectral OCT instrument combined with an air tonometer in a non-collinear configuration was used to image the corneal deformation over full corneal cross-sections, as well as to obtain high speed measurements of the temporal evolution of the corneal apex. The entire deformation process can be dynamically visualized. A quantitative analysis allows direct extraction of several deformation parameters, such as amplitude, diameter and volume of the maximum deformation, as well as duration and speed of the increasing deformation period and the recovery period. The potential of the technique is demonstrated on porcine corneas in vitro under constant IOP for several conditions (untreated, after riboflavin instillation and under cross-linking with ultraviolet light), as well as on human corneas in vivo. The new technique has proved very sensitive to detect differences in the deformation parameters across conditions. We have confirmed non-invasively that Riboflavin and UV-cross-linking induce changes in the corneal biomechanical properties. Those differences appear to be the result of changes in constituent properties of the cornea, and not a consequence of changes in corneal thickness, geometry or IOP. These measurements are a first step for the estimation of the biomechanical properties of corneal tissue, at an individual level and in vivo, to improve diagnosis and prognosis of diseases and treatments involving changes in the biomechanical properties of the cornea. PMID:22435096

Quantified histopathology of the keratoconic cornea.

PubMed

Mathew, Jessica H; Goosey, John D; Bergmanson, Jan P G

2011-08-01

This study systematically investigated and quantified histopathological changes in a series of keratoconic (Kc) corneas using a physiologically formulated fixative to not further distort the already distorted diseased corneas. Twelve surgically removed Kc corneal buttons were immediately preserved and processed for light and transmission electron microscopy using an established corneal protocol. Measurements were taken from the central cone and peripheral regions of the host button. The sample size examined ranged in length from 390 to 2608 μm centrally and 439 to 2242 μm peripherally. The average corneal thickness was 437 μm centrally and 559 μm peripherally. Epithelial thickness varied centrally from 14 to 92 μm and peripherally from 30 to 91 μm. A marked thickening of the epithelial basement membrane was noted in 58% of corneas. Centrally, anterior limiting lamina (ALL) was thinned or lost over 60% of the area examined, whereas peripheral cornea was also affected but to a lesser extent. Histopathologically, posterior cornea remained undisturbed by the disease. Anteriorly in the stroma, an increased number of cells and tissue debris were encountered, and some of these cells were clearly not keratocytes. It is concluded that Kc pathology, at least initially, has a distinct anterior focus involving the epithelium, ALL, and anterior stroma. The epithelium had lost its cellular uniformity and was compromised by the loss or damage to the ALL. The activity of the hitherto unreported recruited stromal cells may be to break down and remove ALL and anterior stromal lamellae, leading to the overall thinning that accompanies this disease.

The influence of Australian eye banking practices on corneal graft survival.

PubMed

Keane, Miriam C; Lowe, Marie T; Coster, Douglas J; Pollock, Graeme A; Williams, Keryn A

2013-08-19

To identify eye banking practices that influence corneal graft survival. Prospective cohort study of records of 19,254 followed corneal grafts in 15160 patients, submitted to the Australian Corneal Graft Registry between May 1985 and July 2012. Influence of corneal preservation method (organ culture, moist pot, Optisol, other); death-to-enucleation, death-to-preservation and enucleation-to-graft times; transportation by air; graft era; and indication for graft on probability of graft survival at most recent follow-up. In multivariate analysis, 919 penetrating grafts performed using corneas transported interstate by air exhibited worse survival than 14,684 grafts performed using corneas retrieved and used locally (hazard ratio [HR], 1.44; 95% CI, 1.21-1.73; P = 0.001). This was also the case for traditional lamellar grafts (64 corneas transported by air and 813 used locally; HR, 1.69; 95% CI, 1.03-2.78; P = 0.038). Indication for graft influenced survival of penetrating grafts (4611 keratoconus, 727 emergency or high-risk, 10,265 other indication; global P < 0.001) and traditional lamellar grafts (65 keratoconus, 212 emergency or high-risk, 600 other indication; global P < 0.001). The preservation medium in which corneas used for traditional lamellar grafts were stored exerted a marginal influence on graft survival (global P = 0.047). Donor corneas transported interstate exhibited poorer survival after transplantation than those retrieved and grafted locally. Higher proportions of emergency procedures involving transported corneas did not account for this difference. Where possible, efforts to avoid transportation of corneal tissue by air freight within Australia may be warranted.

[The cornea: stasis and dynamics].

PubMed

Nishida, Teruo

2008-03-01

The physiological roles of the cornea are to conduct external light into the eye, focus it, together with the lens, onto the retina, and to provide rigidity to the entire eyeball. Good vision thus requires maintenance of the transparency and proper refractive shape of the cornea. Although the cornea appears to be a relatively static structure, dynamic processes operate within and around the cornea at the tissue, cell, and molecular level. In this article, I review the mechanisms responsible for maintenance of corneal homeostasis as well as the development of new modes of treatment for various corneal diseases. I. The static cornea: structure and physiological functions. The cornea is derived from ectoderm, so that it can be considered as transparent skin. It is devoid of blood vessels and manifests the highest sensitivity in the entire body. The surface of the cornea is covered by tear fluid, which serves both as a lubricant and as a conduit for regulatory molecules. The cornea is also supplied with oxygen and various nutrients by the aqueous humor and a loop vascular system in addition to tear fluid. The cornea interacts with its surrounding tissues directly as well as indirectly through tear fluid or aqueous humor, with such interactions playing an important role in the regulation of corneal structure and functions. The resident cells of the cornea-epithelial cells, fibroblasts (keratocytes), and endothelial cells--also engage in mutual interactions through network systems. These interactions as well as those with infiltrated cells and regulation by nerves contribute to the maintenance of the normal structure and functions of the cornea as well as to the repair of corneal injuries. II. The dynamic cornea: maintenance of structure and functions by network systems. Developments in laser and computer technology have allowed observation of the cells and collagen fibers within the cornea. Furthermore, progress in cell and molecular biology has allowed characterization of dynamic network systems-including cell-cell and cell-extracellular matrix interactions as well as cytokines and neural factors-that contribute to the maintenance of corneal transparency and shape. III. Disruption of network systems: persistent corneal epithelial defects and corneal ulcer. Selection of the appropriate treatment for pathologic lesions of the cornea and the accompanying decrease in visual acuity requires localization of the lesion with regard to the epithelium, stroma, or endothelium of the cornea. In certain instances, however, it is not possible to determine the cause of the problem within the cornea. In such cases, the cause of the pathologic lesion and the target for treatment may lie in the surrounding tissues or environment. For example, corneal epithelial wound healing may be delayed, leading to the development of persistent epithelial defects, as a result of disruption of intercellular junctions between epithelial cells, an abnormality of the corneal basement membrane, altered concentrations of various cytokines in tear fluid, a lowered corneal sensation, or allergic reactions in the lid conjunctiva. Loss of corneal epithelial barrier function can further allow inflammatory cytokines present in tear fluid, together with infiltrated cells, to activate keratocytes and elicit excessive degradation of collagen in the stroma, thereby giving rise to corneal ulcer. IV. Development of new drugs for corneal diseases. We have attempted to apply the results of basic scientific research to the development of new drugs for corneal diseases that remain difficult to treat. The process of authorization for new drugs from the Ministry of Health, Labor, and Welfare takes more than two decades, however. The path from the bench to clinical practice is thus a long one. 1. Development of eyedrops for treatment of persistent corneal epithelial defects. We demonstrated the clinical efficacy of fibronectin eyedrops for the treatment of persistent epithelial defects of the cornea. However, the possibility of blood-borne infections has interfered with the development of serum-derived fibronectin as a drug. An automated machine for the preparation of autologous fibronectin eyedrops has therefore recently been developed. Furthermore, in seeking an alternative to fibronectin eyedrops, we are investigating the effects of a peptide corresponding to the second cell-binding domain of fibronectin on corneal epithelial wound healing. Considering that urokinase-type plasminogen activator may be expressed at the site of corneal epithelial defects and facilitates epithelial migration, the potential clinical application of annexin V, which stimulates the secretion of urokinase-type plasminogen activator for the treatment of persistent corneal epithelial defects is also now under investigation in Japan. 2. Development of eyedrops for treatment of neurotrophic keratopathy. Substance P, a neurotransmitter, stimulates corneal epithelial migration in a synergistic manner with insulin-like growth factor (IGF)--1. We have shown that eyedrops containing both the substance P-derived peptide FGLM-amide and the IGF-1--derived peptide SSSR are effective for the treatment of persistent corneal epithelial defects in individuals with diabetic keratopathy or neurotrophic keratopathy, both of which are associated with a reduction in corneal sensation. 3. Development of drugs for corneal ulcer. Treatment of corneal infection with antibiotics does not necessarily halt the process of corneal ulceration, which is characterized by excessive degradation of stromal collagen, or resolve persistent corneal epithelial defects. In addition to eyedrops for the treatment of persistent corneal epithelial defects, we have therefore also been working on the development of new drugs for the treatment of corneal ulcer. To this end, we have established an experimental system in which corneal fibroblasts are cultured in a three-dimensional collagen gel. With this system, we have shown that triptolide and steroids inhibit collagen degradation by corneal fibroblasts. Triptolide or its derivatives are thus potential drugs for the treatment of corneal ulcer and would work by acting directly on corneal fibroblasts rather than by inhibiting the secreted enzymes(matrix metalloproteinases) responsible for collagen degradation.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Skrinnik, A.V.

The author attempts to obtain direct proof of increased permeability of the cornea and enhanced sorption properties of the tissue structures and refractive media of the eye under the influence of a pulsed magnetic field (MF). In the course of the investigation, the method of radioactive indication of two substances (/sup 35/S-streptomycin and /sup 75/Se-methionine), widely used in opthalmologic practice, was used. The radioactivity of the working solution of the preparations was 300,000 cpm in 0.1 ml. Changes in the permeability of the cornea were assessed on the basis of changes in radioactivity of the aqueous humor. The concentration ofmore » /sup 35/S-streptomycin in aqueous humor of the animals immediately after exposure to MF was significantly higher than in the control. Analysis of permeability of the cornea for /sup 75/Se-methionine also showed increased penetration of radioactivity into the aqueous. The results are evidence of the greater penetrating power of /sup 75/Se-methionine.« less

Genetics Home Reference: lattice corneal dystrophy type I

MedlinePlus

... have recurrent corneal erosions, which are caused by separation of particular layers of the cornea from one ... intricate network that forms in the spaces between cells and provides structural support to tissues. The protein ...

Facts about the Cornea and Corneal Disease

MedlinePlus

... can be used as a laboratory model to test potential therapies, or used as therapies on their own to promote regrowth of healthy corneal tissue. For example, one group of researchers is developing a source ...

A new paradigm for use of ultrafast lasers in ophthalmology for enhancement of corneal mechanical properties and permanent correction of refractive errors

NASA Astrophysics Data System (ADS)

Wang, Chao; Fomovsky, Mikhail; Hall, Jamie R.; Paik, David C.; Trokel, Stephen L.; Vukelic, Sinisa

2017-02-01

A new paradigm for strengthening of corneal tissue as well as permanent correction of refractive errors has been proposed. Ultrafast laser irradiation is confined to the levels below optical breakdown such that tissue damage is avoided while creating an ionization field responsible for subsequent photochemical modification of the stroma. The concept was assed using newly developed platform for precise application of a near-IR femtosecond laser irradiation to the cornea in in-vitro experiments. Targeted irradiation with tightly focused ultrafast laser pulses allows spatially resolved crosslinking in the interior of the porcine cornea in the absence of photosensitizers. The formation of intra- or interstromal covalent bonds in collagen matrix locally increases lamellar density. Due to high resolution, treatment is spatially resolved and therefore can be tailored to either enhance structure of corneal stroma or adjust corneal curvature towards correcting refractive errors. As the induced modification is primarily driven by nonlinear absorption, the treatment is essentially wavelength independent, and as such potentially less harmful than current method of choice, joint application of UVA light irradiation in conjunction with riboflavin. Potential applicability of a near-IR femtosecond laser for biomechanical stabilization of cornea and non-invasive refractive eye corrections is discussed.

Impact insertion of transfer-molded microneedle for localized and minimally invasive ocular drug delivery.

PubMed

Song, Hyun Beom; Lee, Kang Ju; Seo, Il Ho; Lee, Ji Yong; Lee, Sang-Mok; Kim, Jin Hyoung; Kim, Jeong Hun; Ryu, WonHyoung

2015-07-10

It has been challenging for microneedles to deliver drugs effectively to thin tissues with little background support such as the cornea. Herein, we designed a microneedle pen system, a single microneedle with a spring-loaded microneedle applicator to provide impact insertion. To firmly attach solid microneedles with 140 μm in height at the end of macro-scale applicators, a transfer molding process was employed. The fabricated microneedle pens were then applied to mouse corneas. The microneedle pens successfully delivered rhodamine dye deep enough to reach the stromal layer of the cornea with small entry only about 1000 μm(2). When compared with syringes or 30 G needle tips, microneedle pens could achieve more localized and minimally invasive delivery without any chances of perforation. To investigate the efficacy of microneedle pens as a way of drug delivery, sunitinib malate proven to inhibit in vitro angiogenesis, was delivered to suture-induced angiogenesis model. When compared with delivery by a 30 G needle tip dipped with sunitinib malate, only delivery by microneedle pens could effectively inhibit corneal neovascularization in vivo. Microneedle pens could effectively deliver drugs to thin tissues without impairing merits of using microneedles: localized and minimally invasive delivery. Copyright © 2015 Elsevier B.V. All rights reserved.

Corneal transplant

MedlinePlus

... and prevent eye movement during the surgery. The tissue for your corneal transplant will come from a person (donor) who has recently died. The donated cornea is processed and tested by a local eye bank to make sure it is safe for use ...

An “All-laser” Endothelial Transplant

PubMed Central

Rossi, Francesca; Canovetti, Annalisa; Malandrini, Alex; Lenzetti, Ivo; Pini, Roberto; Menabuoni, Luca

2015-01-01

The “all laser” assisted endothelial keratoplasty is a procedure that is performed with a femtosecond laser used to cut the donor tissue at an intended depth, and a near infrared diode laser to weld the corneal tissue. The proposed technique enables to reach the three main goals in endothelial keratoplasty: a precise control in the thickness of the donor tissue; its easy insertion in the recipient bed and a reduced risk of donor lenticule dislocation. The donor cornea thickness is measured in the surgery room with optical coherence tomography (OCT), in order to correctly design the donor tissue dimensions. A femtosecond laser is used to cut the donor cornea. The recipient eye is prepared by manual stripping of the descemetic membrane. The donor endothelium is inserted into a Busin-injector, the peripheral inner side is stained with a proper chromophore (a water solution of Indocyanine Green) and then it is pulled in the anterior chamber. The transplanted tissue is placed in the final and correct location and then diode laser welding is induced from outside the eyeball. The procedure has been performed on more than 15 patients evidencing an improvement in surgery performances, with a good recovery of visual acuity and a reduced donor lenticule dislocation event. PMID:26167711

Diffusion and Monod kinetics model to determine in vivo human corneal oxygen-consumption rate during soft contact lens wear.

PubMed

Del Castillo, Luis F; da Silva, Ana R Ferreira; Hernández, Saul I; Aguilella, M; Andrio, Andreu; Mollá, Sergio; Compañ, Vicente

2015-01-01

We present an analysis of the corneal oxygen consumption Qc from non-linear models, using data of oxygen partial pressure or tension (P(O2) ) obtained from in vivo estimation previously reported by other authors. (1) METHODS: Assuming that the cornea is a single homogeneous layer, the oxygen permeability through the cornea will be the same regardless of the type of lens that is available on it. The obtention of the real value of the maximum oxygen consumption rate Qc,max is very important because this parameter is directly related with the gradient pressure profile into the cornea and moreover, the real corneal oxygen consumption is influenced by both anterior and posterior oxygen fluxes. Our calculations give different values for the maximum oxygen consumption rate Qc,max, when different oxygen pressure values (high and low P(O2)) are considered at the interface cornea-tears film. Present results are relevant for the calculation on the partial pressure of oxygen, available at different depths into the corneal tissue behind contact lenses of different oxygen transmissibility. Copyright © 2014. Published by Elsevier Espana.

Raman Spectroscopy of Ocular Tissue

NASA Astrophysics Data System (ADS)

Ermakov, Igor V.; Sharifzadeh, Mohsen; Gellermann, Warner

The optically transparent nature of the human eye has motivated numerous Raman studies aimed at the non-invasive optical probing of ocular tissue components critical to healthy vision. Investigations include the qualitative and quantitative detection of tissue-specific molecular constituents, compositional changes occurring with development of ocular pathology, and the detection and tracking of ocular drugs and nutritional supplements. Motivated by a better understanding of the molecular mechanisms leading to cataract formation in the aging human lens, a great deal of work has centered on the Raman detection of proteins and water content in the lens. Several protein groups and the hydroxyl response are readily detectable. Changes of protein compositions can be studied in excised noncataractous tissue versus aged tissue preparations as well as in tissue samples with artificially induced cataracts. Most of these studies are carried out in vitro using suitable animal models and conventional Raman techniques. Tissue water content plays an important role in optimum light transmission of the outermost transparent ocular structure, the cornea. Using confocal Raman spectroscopy techniques, it has been possible to non-invasively measure the water to protein ratio as a measure of hydration status and to track drug-induced changes of the hydration levels in the rabbit cornea at various depths. The aqueous humor, normally supplying nutrients to cornea and lens, has an advantageous anterior location for Raman studies. Increasing efforts are pursued to non-invasively detect the presence of glucose and therapeutic concentrations of antibiotic drugs in this medium. In retinal tissue, Raman spectroscopy proves to be an important tool for research into the causes of macular degeneration, the leading cause of irreversible vision disorders and blindness in the elderly. It has been possible to detect the spectral features of advanced glycation and advanced lipooxydation end products in excised tissue samples and synthetic preparations and thus to identify potential biomarkers for the onset of this disease. Using resonance Raman detection techniques, the concentration and spatial distribution of macular pigment, a protective compound, can be detected in the living human retina Useable in clinical settings for patient screening, the technology is suitable to investigate correlations between pigment concentration levels and risk for macular degeneration and to monitor increases in pigment levels occurring as a result of dietary intervention strategies.

Synthetic cornea: biocompatibility and optics

NASA Astrophysics Data System (ADS)

Parel, Jean-Marie A.; Kaminski, Stefan; Fernandez, Viviana; Alfonso, E.; Lamar, Peggy; Lacombe, Emmanuel; Duchesne, Bernard; Dubovy, Sander; Manns, Fabrice; Rol, Pascal O.

2002-06-01

Purpose. Experimentally find a method to provide a safe surgical technique and an inexpensive and long lasting mesoplant for the restoration of vision in patients with bilateral corneal blindness due to ocular surface and stromal diseases. Methods. Identify the least invasive and the safest surgical technique for synthetic cornea implantation. Identify the most compatible biomaterials and the optimal shape a synthetic cornea must have to last a long time when implanted in vivo. Results. Penetrating procedures were deemed too invasive, time consuming, difficult and prone to long term complications. Therefore a non-penetrating delamination technique with central trephination was developed to preserve the integrity of Descemet's membrane and the anterior segment. Even though this approach limits the number of indications, it is acceptable since the majority of patients only have opacities in the stroma. The prosthesis was designed to fit in the removed tissue plane with its skirt fitted under the delaminated stroma. To improve retention, the trephination wall was made conical with the smallest opening on the anterior surface and a hat-shaped mesoplant was made to fit. The skirt was perforated in its perimeter to allow passage of nutrients and tissues ingrowths. To simplify the fabrication procedure, the haptic and optic were made of the same polymer. The intrastromal biocompatibility of several hydrogels was found superior to current clinically used PMMA and PTFE materials. Monobloc mesoplants made of 4 different materials were implanted in rabbits and followed weekly until extrusion occurred. Some remained optically clear allowing for fundus photography. Conclusions. Hydrogel synthetic corneas can be made to survive for periods longer than 1 year. ArF excimer laser photoablation studies are needed to determine the refractive correction potential of these mesoplants. A pilot FDA clinical trial is needed to assess the mesoplant efficacy and very long-term stability.

Changes in the Refractive Index of the Stroma and Its Extrafibrillar Matrix When the Cornea Swells

PubMed Central

Meek, Keith M.; Dennis, Sally; Khan, Shukria

2003-01-01

The transparency of the corneal stroma is critically dependent on the hydration of the tissue; if the cornea swells, light scattering increases. Although this scattering has been ascribed to the disruption caused to the arrangement of the collagen fibrils, theory predicts that light scattering could increase if there is an increased mismatch in the refractive indices of the collagen fibrils and the material between them. The purpose of this article is to use Gladstone and Dale's law of mixtures to calculate volume fractions for a number of different constituents in the stroma, and use these to show how the refractive indices of the stroma and its constituent extrafibrillar material would be expected to change as more solvent enters the tissue. Our calculations predict that solvent entering the extrafibrillar space causes a reduction in its refractive index, and hence a reduction in the overall refractive index of the bovine stroma according to the equation n′s = 1.335 + 0.04/(0.22 + 0.24 H′), where n′s is the refractive index and H′ is the hydration of the swollen stroma. This expression is in reasonable agreement with our experimental measurements of refractive index versus hydration in bovine corneas. When the hydration of the stroma increases from H = 3.2 to H = 8.0, we predict that the ratio of the refractive index of the collagen fibrils to that of the material between them increases from 1.041 to 1.052. This change would be expected to make only a small contribution to the large increase in light scattering observed when the cornea swells to H = 8. PMID:14507686

Donor cross-linking for keratoplasty: a laboratory evaluation.

PubMed

Mukherjee, Achyut; Hayes, Sally; Aslanides, Ioannis; Lanchares, Elena; Meek, Keith M

2015-12-01

This laboratory-based investigation compares the topographic outcomes of conventional penetrating keratoplasty with that of a novel procedure in which donor corneas are cross-linked prior to keratoplasty. Penetrating keratoplasty procedures with continuous running sutures were carried out in a porcine whole globe model. Sixty eyes were randomly paired as 'donor' and 'host' tissue before being assigned to one of two groups. In the cross-linked group, donor corneas underwent riboflavin/UVA cross-linking prior to being trephined and sutured to untreated hosts. In the conventional keratoplasty group, both host and donor corneas remained untreated prior to keratoplasty. Topographic and corneal wavefront measurements were performed following surgery, and technical aspects of the procedure evaluated. Mean keratometric astigmatism was significantly lower in the cross-linked donor group at 3.67D (SD 1.8 D), vs. 8.43 D (SD 2.4 D) in the conventional keratoplasty group (p < 0.005). Mean wavefront astigmatism was also significantly reduced in the cross-linked donor group 4.71 D (SD 2.1) vs. 8.29D (SD 3.6) in the conventional keratoplasty group (p < 0.005). Mean RMS higher order aberration was significantly lower in the cross-linked donor group at 1.79 um (SD 0.98), vs. 3.05 um (SD 1.9) in the conventional keratoplasty group (P = 0.02). Qualitative analysis revealed less tissue distortion at the graft-host junction in the cross-linked group. Cross-linking of donor corneas prior to keratoplasty reduces intraoperative induced astigmatism and aberrations in an animal model. Further studies are indicated to evaluate the implications of this potential modification of keratoplasty surgery.

Alternatives to eye bank native tissue for corneal stromal replacement.

PubMed

Brunette, Isabelle; Roberts, Cynthia J; Vidal, François; Harissi-Dagher, Mona; Lachaine, Jean; Sheardown, Heather; Durr, Georges M; Proulx, Stéphanie; Griffith, May

2017-07-01

Corneal blindness is a major cause of blindness in the world and corneal transplantation is the only widely accepted treatment to restore sight in these eyes. However, it is becoming increasingly difficult for eye banks to meet the increasing demand for transplantable tissue, which is in part due to population aging. Donor tissue shortage is therefore a growing concern globally and there is a need for alternatives to human donor corneas. Biosynthetic corneal substitutes offer several significant advantages over native corneas: Large-scale production offers a powerful potential solution to the severe shortage of human donor corneas worldwide; Good manufacturing practices ensure sterility and quality control; Acellular corneal substitutes circumvent immune rejection induced by allogeneic cells; Optical and biomechanical properties of the implants can be adapted to the clinical need; and finally these corneal substitutes could benefit from new advances in biomaterials science, such as surface coating, functionalization and nanoparticles. This review highlights critical contributions from laboratories working on corneal stromal substitutes. It focuses on synthetic inert prostheses (keratoprostheses), acellular scaffolds with and without enhancement of endogenous regeneration, and cell-based replacements. Accent is put on the physical properties and biocompatibility of these biomaterials, on the functional and clinical outcome once transplanted in vivo in animal or human eyes, as well as on the main challenges of corneal stromal replacement. Regulatory and economic aspects are also discussed. All of these perspectives combined highlight the founding principles of the clinical application of corneal stromal replacement, a concept that has now become reality. Copyright © 2017 Elsevier Ltd. All rights reserved.

Membrane nanotubes in myeloid cells in the adult mouse cornea represent a novel mode of immune cell interaction.

PubMed

Seyed-Razavi, Yashar; Hickey, Michael J; Kuffová, Lucia; McMenamin, Paul G; Chinnery, Holly R

2013-01-01

Membrane nanotubes (MNTs) are newly discovered cellular extensions that are either blind-ended or can connect widely separated cells. They have predominantly been investigated in cultured isolated cells, however, previously we were the first group to demonstrate the existence of these structures in vivo in intact mammalian tissues. We previously demonstrated the frequency of both cell-cell or bridging MNTs and blind-ended MNTs was greatest between major histocompatibility complex (MHC) class II(+) cells during corneal injury or TLR ligand-mediated inflammation. The present study aimed to further explore the dynamics of MNT formation and their size, presence in another tissue, the dura mater, and response to stress factors and an active local viral infection of the murine cornea. Confocal live cell imaging of myeloid-derived cells in inflamed corneal explants from Cx(3)cr1(GFP) and CD11c(eYFP) transgenic mice revealed that MNTs form de novo at a rate of 15.5 μm/min. This observation contrasts with previous studies that demonstrated that in vitro these structures originate from cell-cell contacts. Conditions that promote formation of MNTs include inflammation in vivo and cell stress due to serum starvation ex vivo. Herpes simplex virus-1 infection did not cause a significant increase in MNT numbers in myeloid cells in the cornea above that observed in injury controls, confirming that corneal epithelium injury alone elicits MNT formation in vivo. These novel observations extend the currently limited understanding of MNTs in live mammalian tissues.

Infrared laser damage thresholds in corneal tissue phantoms using femtosecond laser pulses

NASA Astrophysics Data System (ADS)

Boretsky, Adam R.; Clary, Joseph E.; Noojin, Gary D.; Rockwell, Benjamin A.

2018-02-01

Ultrafast lasers have become a fixture in many biomedical, industrial, telecommunications, and defense applications in recent years. These sources are capable of generating extremely high peak power that can cause laser-induced tissue breakdown through the formation of a plasma upon exposure. Despite the increasing prevalence of such lasers, current safety standards (ANSI Z136.1-2014) do not include maximum permissible exposure (MPE) values for the cornea with pulse durations less than one nanosecond. This study was designed to measure damage thresholds in corneal tissue phantoms in the near-infrared and mid-infrared to identify the wavelength dependence of laser damage thresholds from 1200-2500 nm. A high-energy regenerative amplifier and optical parametric amplifier outputting 100 femtosecond pulses with pulse energies up to 2 mJ were used to perform exposures and determine damage thresholds in transparent collagen gel tissue phantoms. Three-dimensional imaging, primarily optical coherence tomography, was used to evaluate tissue phantoms following exposure to determine ablation characteristics at the surface and within the bulk material. The determination of laser damage thresholds in the near-IR and mid-IR for ultrafast lasers will help to guide safety standards and establish the appropriate MPE levels for exposure sensitive ocular tissue such as the cornea. These data will help promote the safe use of ultrafast lasers for a wide range of applications.

Density of ocular components of the bovine eye.

PubMed

Su, Xiao; Vesco, Christina; Fleming, Jacquelyn; Choh, Vivian

2009-10-01

Density is essential for acoustic characterization of tissues and provides a basic input for ultrasound backscatter and absorption models. Despite the existence of extensive compilations of acoustic properties, neither unified data on ocular density nor comparisons of the densities between all ocular components can be found. This study was undertaken to determine the mass density of all the ocular components of the bovine eye. Liquid components were measured through mass/volume ratio, whereas solid tissues were measured with two different densitometry techniques based on Archimedes Principle. The first method determines the density by measuring dry and wet weight of the tissues. The second method consists of immersing the tissues in sucrose solutions of varying densities and observing their buoyancy. Although the mean densities for all tissues were found to be within 0.02 g/cm by both methods, only the sucrose solution method offered a consistent relative order for all measured ocular components, as well as a considerably smaller standard deviation (a maximum standard deviation of 0.004 g/cm for cornea). The lens was found to be the densest component, followed by the sclera, cornea, choroid, retina, aqueous, and vitreous humors. The consistent results of the sucrose solution tests suggest that the ocular mass density is a physical property that is more dependent on the compositional and structural characteristics of the tissue and than on population variability.

Brillouin microscopy: assessing ocular tissue biomechanics.

PubMed

Yun, Seok Hyun; Chernyak, Dimitri

2018-07-01

Assessment of corneal biomechanics has been an unmet clinical need in ophthalmology for many years. Many researchers and clinicians have identified corneal biomechanics as source of variability in refractive procedures and one of the main factors in keratoconus. However, it has been difficult to accurately characterize corneal biomechanics in patients. The recent development of Brillouin light scattering microscopy heightens the promise of bringing biomechanics into the clinic. The aim of this review is to overview the progress and discuss prospective applications of this new technology. Brillouin microscopy uses a low-power near-infrared laser beam to determine longitudinal modulus or mechanical compressibility of tissue by analyzing the return signal spectrum. Human clinical studies have demonstrated significant difference in the elastic properties of normal corneas versus corneas diagnosed with mild and severe keratoconus. Clinical data have also shown biomechanical changes after corneal cross-linking treatment of keratoconus patients. Brillouin measurements of the crystalline lens and sclera have also been demonstrated. Brillouin microscopy is a promising technology under commercial development at present. The technique enables physicians to characterize the biomechanical properties of ocular tissues.

Cell-penetrating peptide for enhanced delivery of nucleic acids and drugs to ocular tissues including retina and cornea.

PubMed

Johnson, Leslie N; Cashman, Siobhan M; Kumar-Singh, Rajendra

2008-01-01

As in other organ systems, gene and drug delivery to ocular tissues such as the retina and cornea is hampered by inefficient penetration of therapeutic molecules across the plasma membrane. We describe the use of a novel peptide for ocular delivery (POD) with protein transduction properties, for delivery of small and large molecules across the plasma membrane. POD enters cells within 5 minutes in a temperature dependent manner. POD can compact and deliver plasmid DNA, achieving transgene expression in >50% of human embryonic retinoblasts. Delivery of small interfering RNA (siRNA) duplexes to cells using POD, allowed for silencing of transgene expression by >50%. POD could also be used to deliver quantum dots in vitro and in vivo. Upon ocular delivery, POD rapidly entered neural retina and localized to retinal pigment epithelium (RPE), photoreceptor, and ganglion cells. Additionally, POD was able to enter corneal epithelium, sclera, choroid, and the dura of the optic nerve via topical application. POD also functions as a bacteriostatic, a useful property for a carrier of molecules to post mitotic neural ocular tissues.

Chronic Lunar Dust Exposure on Rat Cornea: Evaluation by Gene Expression Profiling

NASA Technical Reports Server (NTRS)

Theriot, C. A.; Glass, A.; Lam, C-W.; James, J.; Zanello, S. B.

2014-01-01

Lunar dust is capable of entering habitats and vehicle compartments by sticking to spacesuits or other objects that are transferred into the spacecraft from the lunar surface and has been reported to cause irritation upon exposure. During the Apollo missions, crewmembers reported irritation specifically to the skin and eyes after contamination of the lunar and service modules. It has since been hypothesized that ocular irritation and abrasion might occur as a result of such exposure, impairing crew vision. Recent work has shown that both ultrafine and unground lunar dust exhibited minimal irritancy of the ocular surface (i.e., cornea); however, the assessment of the severity of ocular damage resulting from contact of lunar dust particles to the cornea has focused only on macroscopic signs of mechanical irritancy and cytotoxicity. Given the chemical reactive properties of lunar dust, exposure of the cornea may contribute to detrimental effects at the molecular level including but not limited to oxidative damage. Additionally, low level chronic exposures may confound any results obtained in previous acute studies. We report here preliminary results from a tissue sharing effort using 10-week-old Fischer 344 male rats chronically exposed to filtered air or jet milled lunar dust collected during Apollo 14 using a Jaeger-NYU nose-only chamber for a total of 120 hours (6 hours daily, 5 days a week) over a 4-week period. RNA was isolated from corneas collected from rats at 1 day and 7 days after being exposed to concentrations of 0, 20, and 60 mg/m3 of lunar dust. Microarray analysis was performed using the Affymetrix GeneChip Rat Genome 230 2.0 Array with Affymetrix Expression Console and Transcriptome Analysis Console used for normalization and secondary analysis. An Ingenuity iReport"TM" was then generated for canonical pathway identification. The number of differentially expressed genes identified increases with dose compared to controls suggesting a more severe response to the lunar dust insult at higher levels. Pathways of interests that have been identified in all exposed samples include oxidative stress response, mitochondrial dysfunction, fibrosis, epithelial healing, TGF-Beta? signaling, and extracellular matrix remodeling. Several biological processes related to cell migration, cellular proliferation, and eye development were also identified to be altered by exposure to lunar dust. Our preliminary results suggest that even a chronic insult of lunar dust as low as 20 mg/m(exp 3) elicits a molecular response in cornea tissue. Lunar dust on the surface of the moon would have the added properties of ionization and activation potentially leading to further damage to the cornea and greater sensitivity to any other environmental insult such as exposure to radiation. Additional studies are required to fully assess the risk of vision impairment and the mechanistic responses initiated in cornea exposed to lunar dust as well as the potential for long-term effects to astronaut health

Optimization of immunostaining on flat-mounted human corneas.

PubMed

Forest, Fabien; Thuret, Gilles; Gain, Philippe; Dumollard, Jean-Marc; Peoc'h, Michel; Perrache, Chantal; He, Zhiguo

2015-01-01

In the literature, immunohistochemistry on cross sections is the main technique used to study protein expression in corneal endothelial cells (ECs), even though this method allows visualization of few ECs, without clear subcellular localization, and is subject to the staining artifacts frequently encountered at tissue borders. We previously proposed several protocols, using fixation in 0.5% paraformaldehyde (PFA) or in methanol, allowing immunostaining on flatmounted corneas for proteins of different cell compartments. In the present study, we further refined the technique by systematically assessing the effect of fixative temperature. Last, we used optimized protocols to further demonstrate the considerable advantages of immunostaining on flatmounted intact corneas: detection of rare cells in large fields of thousands of ECs and epithelial cells, and accurate subcellular localization of given proteins. The staining of four ubiquitous proteins, ZO-1, hnRNP L, actin, and histone H3, with clearly different subcellular localizations, was analyzed in ECs of organ-cultured corneas. Whole intact human corneas were fixed for 30 min in 0.5% paraformaldehyde or pure methanol at four temperatures (4 °C for PFA, -20 °C for methanol, and 23, 37, and 50 °C for both). Experiments were performed in duplicate and repeated on three corneas. Standardized pictures were analyzed independently by two experts. Second, optimized immunostaining protocols were applied to fresh corneas for three applications: identification of rare cells that express KI67 in the endothelium of specimens with Fuch's endothelial corneal dystrophy (FECD), the precise localization of neural cell adhesion molecules (NCAMs) in normal ECs and of the cytokeratin pair K3/12 and CD44 in normal epithelial cells, and the identification of cells that express S100b in the normal epithelium. Temperature strongly influenced immunostaining quality. There was no ubiquitous protocol, but nevertheless, room temperature may be recommended as first-line temperature during fixation, instead of the conventional -20 °C for methanol and 4 °C for PFA. Further optimization may be required for certain target proteins. Optimized protocols allowed description of two previously unknown findings: the presence of a few proliferating ECs in FECD specimens, suggesting ineffective compensatory mechanisms against premature EC death, and the localization of NCAMs exclusively in the lateral membranes of ECs, showing hexagonal organization at the apical pole and an irregular shape with increasing complexity toward the basal pole. Optimized protocols were also effective for the epithelium, allowing clear localization of cytokeratin 3/12 and CD44 in superficial and basal epithelial cells, respectively. Finally, S100b allowed identification of clusters of epithelial Langerhans cells near the limbus and more centrally. Fixative temperature is a crucial parameter in optimizing immunostaining on flatmounted intact corneas. Whole-tissue overview and precise subcellular staining are significant advantages over conventional immunohistochemistry (IHC) on cross sections. This technique, initially developed for the corneal endothelium, proved equally suitable for the corneal epithelium and could be used for other superficial mono- and multilayered epithelia.

Infrared irradiation alters the expression of matrix metalloproteinases and glycosaminoglycans in the cornea and crystalline lens.

PubMed

Dadoukis, Panagiotis; Klagas, Ioannis; Komnenou, Anastasia; Karakiulakis, George; Karoutis, Athanasios; Karampatakis, Vassilios; Papakonstantinou, Eleni

2013-08-01

Prolonged exposure to infrared (IR) radiation is associated with different types of damage to cornea and lens. The aim of our study was to investigate the effect of acute and chronic exposure to IR radiation on the activity of matrix metalloproteinase-2 (MMP-2) and MMP-9 and on the expression of glycosaminoglycans (GAG) in the rabbit cornea and crystalline lens. New Zealand rabbits were subjected to IR radiation for 4 months (chronic exposure to IR) or to normal light (control group). In experiments regarding acute exposure, animals were subjected to IR radiation or normal light for 12 h, in the presence of 0.1% diclofenac sodium (eye drops instilled in the right eye of animals) or saline (instilled in the left eye of animals). The cornea and lens were dissected away and homogenized. The activity of MMP-2 and MMP-9 was assayed by gelatine zymography. Total GAG were isolated from tissue specimens after lipid extraction and extensive digestion with pronase and DNase and characterized by treatment with GAG-degrading enzymes, followed by electrophoresis on cellulose acetate membranes. Acute or chronic exposure to IR radiation induced the activity of MMP-2 in cornea and lens, whereas only acute IR radiation increased the content of heparan sulphate in crystalline lens. Local administration of diclofenac sodium did not prevent the above effects of acute IR radiation. The detrimental effects of excessive or prolonged exposure of the eyes to IR radiation are associated with induced activity of MMP-2 in cornea and lens and alterations in the content of heparan sulphate in lens. Thus, MMP and GAG may offer alternative targets for pharmacological intervention to confront ocular damages associated with IR radiation.

Noninvasive intratissue refractive index shaping (IRIS) of the cornea with blue femtosecond laser light.

PubMed

Xu, Lisen; Knox, Wayne H; DeMagistris, Margaret; Wang, Nadan; Huxlin, Krystel R

2011-10-17

To test the feasibility of intratissue refractive index shaping (IRIS) in living corneas by using 400-nm femtosecond (fs) laser pulses (blue-IRIS). To test the hypothesis that the intrinsic two-photon absorption of the cornea allows blue-IRIS to be performed with greater efficacy than when using 800-nm femtosecond laser pulses. Fresh cat corneas were obtained postmortem and cut into six wedges. Blue laser pulses at 400 nm, with 100-fs pulse duration at 80 MHz were used to micromachine phase gratings into each corneal wedge at scanning speeds from 1 to 15 mm/s. Grating lines were 1 μm wide, 5 μm apart, and 150 μm below the anterior corneal surface. Refractive index (RI) changes in micromachined regions were measured immediately by recording the diffraction efficiency of inscribed gratings. Six hours later, the corneas were processed for histology, and TUNEL staining was performed to assess whether blue-IRIS causes cell death. Scanning at 1 and 2 mm/s caused overt corneal damage in the form of bubbles and burns. At faster scanning speeds (5, 10, and 15 mm/s), phase gratings were created in the corneal stroma, which were shown to be pure RI changes ranging from 0.037 to 0.021 in magnitude. The magnitude of RI change was inversely related to scanning speed. TUNEL staining showed cell death only around bubbles and burns. Blue-IRIS can be performed safely and effectively in living cornea. Compared with near-infrared laser pulses, blue-IRIS enhances both achievable RI change and scanning speed without the need to dope the tissue with two-photon sensitizers, increasing the clinical applicability of this technique.

Noninvasive Intratissue Refractive Index Shaping (IRIS) of the Cornea with Blue Femtosecond Laser Light

PubMed Central

Xu, Lisen; Knox, Wayne H.; DeMagistris, Margaret; Wang, Nadan

2011-01-01

Purpose. To test the feasibility of intratissue refractive index shaping (IRIS) in living corneas by using 400-nm femtosecond (fs) laser pulses (blue-IRIS). To test the hypothesis that the intrinsic two-photon absorption of the cornea allows blue-IRIS to be performed with greater efficacy than when using 800-nm femtosecond laser pulses. Methods. Fresh cat corneas were obtained postmortem and cut into six wedges. Blue laser pulses at 400 nm, with 100-fs pulse duration at 80 MHz were used to micromachine phase gratings into each corneal wedge at scanning speeds from 1 to 15 mm/s. Grating lines were 1 μm wide, 5 μm apart, and 150 μm below the anterior corneal surface. Refractive index (RI) changes in micromachined regions were measured immediately by recording the diffraction efficiency of inscribed gratings. Six hours later, the corneas were processed for histology, and TUNEL staining was performed to assess whether blue-IRIS causes cell death. Results. Scanning at 1 and 2 mm/s caused overt corneal damage in the form of bubbles and burns. At faster scanning speeds (5, 10, and 15 mm/s), phase gratings were created in the corneal stroma, which were shown to be pure RI changes ranging from 0.037 to 0.021 in magnitude. The magnitude of RI change was inversely related to scanning speed. TUNEL staining showed cell death only around bubbles and burns. Conclusions. Blue-IRIS can be performed safely and effectively in living cornea. Compared with near-infrared laser pulses, blue-IRIS enhances both achievable RI change and scanning speed without the need to dope the tissue with two-photon sensitizers, increasing the clinical applicability of this technique. PMID:21931133

Genetic evidence for the coordinated regulation of collagen fibrillogenesis in the cornea by decorin and biglycan.

PubMed

Zhang, Guiyun; Chen, Shoujun; Goldoni, Silvia; Calder, Bennett W; Simpson, Holly C; Owens, Rick T; McQuillan, David J; Young, Marian F; Iozzo, Renato V; Birk, David E

2009-03-27

Decorin and biglycan are class I small leucine-rich proteoglycans (SLRPs) involved in regulation of collagen fibril and matrix assembly. We hypothesize that tissue-specific matrix assembly, such as in the cornea, requires a coordinate regulation involving multiple SLRPs. To this end, we investigated the expression of decorin and biglycan in the cornea of mice deficient in either SLRP gene and in double-mutant mice. Decorin and biglycan exhibited overlapping spatial expression patterns throughout the corneal stroma with differential temporal expression. Whereas decorin was expressed at relatively high levels in all developmental stages, biglycan expression was high early, decreased during development, and was present at very low levels in the mature cornea. Ultrastructural analyses demonstrated comparable fibril structure in the decorin- and biglycan-null corneas compared with wild-type controls. We found a compensatory up-regulation of biglycan gene expression in the decorin-deficient mice, but not the reverse. Notably, the corneas of compound decorin/biglycan-null mice showed severe disruption in fibril structure and organization, especially affecting the posterior corneal regions, corroborating the idea that biglycan compensates for the loss of decorin. Fibrillogenesis assays using recombinant decorin and biglycan confirmed a functional compensation, with both having similar effects at high SLRP/collagen ratios. However, at low ratios decorin was a more efficient regulator. The use of proteoglycan or protein core yielded comparable results. These findings provide firm genetic evidence for an interaction of decorin and biglycan during corneal development and further suggest that decorin has a primary role in regulating fibril assembly, a function that can be fine-tuned by biglycan during early development.

Preliminary observations on the effects of selenate on the development of the embryonic skate, Raja eglanteria

NASA Technical Reports Server (NTRS)

Conrad, G. W.; Luer, C. A.; Paulsen, A. Q.; Funderburgh, J. L.; Spooner, B. S. (Principal Investigator)

1993-01-01

Morphogenesis of the clearnose skate, Raja eglanteria, was not significantly inhibited as a result of 7 days of exposure to 1-2 mM selenate in the sea water during Days 59-69 of embryonic development (hatching would normally have occurred at 82 +/- 4 days of incubation). Although corneal transparency appeared normal in the eye, preliminary measurements of the thickness of Bowman's layer of the cornea suggested that it was significantly thinner in the corneas of embryos exposed to 1-2 mM selenate. Selenate is an ion reported to inhibit sulfation of glycosaminoglycans in connective tissue.

[Corneal manifestations in systemic diseases].

PubMed

Zarranz Ventura, J; De Nova, E; Moreno-Montañés, J

2008-01-01

Systemic diseases affecting the cornea have a wide range of manifestations. The detailed study of all pathologies that cause corneal alteration is unapproachable, so we have centered our interest in the most prevalent or characteristic of them. In this paper we have divided these pathologies in sections to facilitate their study. Pulmonar and conective tissue (like colagen, rheumatologic and idiopathic inflamatory diseases), dermatologic, cardiovascular, hematologic, digestive and hepatopancreatic diseases with corneal alteration are described. Endocrine and metabolic diseases, malnutrition and carential states are also studied, as well as some otorhinolaryngologic and genetic diseases that affect the cornea. Finally, a brief report of ocular toxicity induced by drugs is referred.

West Nile Virus Infection in Human and Mouse Cornea Tissue

PubMed Central

Blitvich, Bradley J.; Wang, Tian; Saxena, Vandana; Zeng, Shemin; Harmon, Karen M.; Raymond, Matthew D.; Goins, Kenneth M.; Reed, Cynthia R.; Mullins, Robert F.; Greiner, Mark A.

2016-01-01

The purpose of this study was to determine the in vitro and ex vivo susceptibility of human corneal cells to West Nile virus (WNV) infection and evaluate the ability of the virus to disseminate to the corneas of infected mice. Human corneal epithelial cells were challenged with WNV, incubated for 1–6 days, and tested for evidence of WNV infection. Viral RNA and antigen were detected at every time point, and the virus reached a peak titer of 2.5 × 107 plaque-forming units (pfu)/mL at 3 days postinoculation (PI). Corneas procured from donors were incubated in culture dishes containing WNV for 1–5 days and tested for evidence of WNV. Viral RNA and antigen were detected, and the virus reached a mean peak titer of 4.9 × 104 pfu/mL at 5 days PI. Mice were inoculated intraperitoneally with WNV, and their eyes were harvested at 2, 5, and 8 days PI and tested for evidence of WNV. Viral RNA was detected in corneas of four of nine systemically infected mice as early as 2 days PI. We conclude that human corneal cells support WNV replication in vitro and ex vivo, and WNV may disseminate into the corneas of experimentally infected mice. These findings indicate that corneal transmission cannot be ruled out as a novel mode of human-to-human WNV transmission and additional experiments should be conducted to assess this risk further. PMID:27672204

The Influence of Lamellar Orientation on Corneal Material Behavior: Biomechanical and Structural Changes in an Avian Corneal Disorder

PubMed Central

Elsheikh, Ahmed; Kassem, Wael; Kamma-Lorger, Christina S.; Hocking, Paul M.; White, Nick; Inglehearn, Chris F.; Ali, Manir; Meek, Keith M.

2011-01-01

Purpose. Retinopathy, globe enlarged (RGE) is an inherited genetic disease of chickens with a corneal phenotype characterized by loss of tissue curvature and changes in peripheral collagen fibril alignment. This study aimed to characterize the material behavior of normal and RGE chicken corneas under inflation and compare this with new spatial- and depth-resolved microstructural information to investigate how stromal fibril architecture determines corneal behavior under intraocular pressure (IOP). Methods. Six RGE chicken corneas and six age-matched normal controls were tested using trephinate inflation and their stress-strain behavior determined as a function of posterior pressure. Second harmonic generation mulitphoton microscopy was used to compare the in-plane appearance and degree of through-plane interlacing of collagen lamellae between normal and mutant corneas. Results. RGE corneas displayed a 30–130% increase in material stiffness [Etangent(RGE) = 0.94 ± 0.18 MPa to 3.09 ± 0.66 MPa; Etangent(normals) = 0.72 ± 0.13 MPa to 1.34 ± 0.35 MPa] (P ≤ 0.05). The normal in-plane disposition of anterior collagen in the peripheral cornea was altered in RGE but through-plane lamellar interlacing was unaffected. Conclusions. This article demonstrates changes in corneal material behavior in RGE that are qualitatively consistent with microstructural collagen alterations identified both herein and previously. This study indicates that, in general, changes in stromal fibril orientation may significantly affect corneal material behavior and thereby its response to IOP. PMID:21051696

West Nile Virus Infection in Human and Mouse Cornea Tissue.

PubMed

Blitvich, Bradley J; Wang, Tian; Saxena, Vandana; Zeng, Shemin; Harmon, Karen M; Raymond, Matthew D; Goins, Kenneth M; Reed, Cynthia R; Mullins, Robert F; Greiner, Mark A

2016-11-02

The purpose of this study was to determine the in vitro and ex vivo susceptibility of human corneal cells to West Nile virus (WNV) infection and evaluate the ability of the virus to disseminate to the corneas of infected mice. Human corneal epithelial cells were challenged with WNV, incubated for 1-6 days, and tested for evidence of WNV infection. Viral RNA and antigen were detected at every time point, and the virus reached a peak titer of 2.5 × 10 7 plaque-forming units (pfu)/mL at 3 days postinoculation (PI). Corneas procured from donors were incubated in culture dishes containing WNV for 1-5 days and tested for evidence of WNV. Viral RNA and antigen were detected, and the virus reached a mean peak titer of 4.9 × 10 4 pfu/mL at 5 days PI. Mice were inoculated intraperitoneally with WNV, and their eyes were harvested at 2, 5, and 8 days PI and tested for evidence of WNV. Viral RNA was detected in corneas of four of nine systemically infected mice as early as 2 days PI. We conclude that human corneal cells support WNV replication in vitro and ex vivo, and WNV may disseminate into the corneas of experimentally infected mice. These findings indicate that corneal transmission cannot be ruled out as a novel mode of human-to-human WNV transmission and additional experiments should be conducted to assess this risk further. © The American Society of Tropical Medicine and Hygiene.

Characterisation of the metabolome of ocular tissues and post-mortem changes in the rat retina.

PubMed

Tan, Shi Z; Mullard, Graham; Hollywood, Katherine A; Dunn, Warwick B; Bishop, Paul N

2016-08-01

Time-dependent post-mortem biochemical changes have been demonstrated in donor cornea and vitreous, but there have been no published studies to date that objectively measure post-mortem changes in the retinal metabolome over time. The aim of the study was firstly, to investigate post-mortem, time-dependent changes in the rat retinal metabolome and secondly, to compare the metabolite composition of healthy rat ocular tissues. To study post-mortem changes in the rat retinal metabolome, globes were enucleated and stored at 4 °C and sampled at 0, 2, 4, 8, 24 and 48 h post-mortem. To study the metabolite composition of rat ocular tissues, eyes were dissected immediately after culling to isolate the cornea, lens, vitreous and retina, prior to storing at -80 °C. Tissue extracts were subjected to Gas Chromatograph Mass Spectrometry (GC-MS) and Ultra High Performance Liquid Chromatography Mass Spectrometry (UHPLC-MS). Generally, the metabolic composition of the retina was stable for 8 h post-mortem when eyes were stored at 4 °C, but showed increasing changes thereafter. However, some more rapid changes were observed such as increases in TCA cycle metabolites after 2 h post-mortem, whereas some metabolites such as fatty acids only showed decreases in concentration from 24 h. A total of 42 metabolites were identified across the ocular tissues by GC-MS (MSI level 1) and 2782 metabolites were annotated by UHPLC-MS (MSI level 2) according to MSI reporting standards. Many of the metabolites detected were common to all of the tissues but some metabolites showed partitioning between different ocular structures with 655, 297, 93 and 13 metabolites being uniquely detected in the retina, lens, cornea and vitreous respectively. Only a small percentage (1.6%) of metabolites found in the vitreous were only detected in the retina and not other tissues. In conclusion, mass spectrometry-based techniques have been used for the first time to compare the metabolic composition of different ocular tissues. The metabolite composition of the retina stored at 4 °C post-mortem is mostly stable for at least 8 h. Copyright © 2016 Elsevier Ltd. All rights reserved.

Delay of corneal epithelial wound healing and induction of keratocyte apoptosis by platelet-activating factor.

PubMed

Chandrasekher, Gudiseva; Ma, Xiang; Lallier, Thomas E; Bazan, Haydee E P

2002-05-01

To examine the role of the lipid mediator platelet-activating factor (PAF) in epithelial wound healing. A 7-mm central de-epithelializing wound was produced in rabbit corneas, and the tissue was incubated with 125 nM carbamyl PAF (cPAF), an analogue of PAF. Rabbit corneal epithelial and stromal cells were also cultured in the presence of cPAF. Cell adhesion, proliferation, and migration assays were conducted. Apoptosis was assayed by TUNEL staining on preparations of corneal tissue sections and in cells in culture. Twenty-four hours after injury, 50% of the wounded area was covered by new epithelium, whereas only 30% was covered in the presence of cPAF. At 48 hours, the epithelium completely closed the wound, but only 45% of the original wound was covered in corneas treated with cPAF. Similar inhibition of epithelial wound closure was found with human corneas incubated with PAF in organ culture. Moreover, addition of several growth factors involved in corneal wound healing, such as epidermal growth factor, hepatocyte growth factor, and keratinocyte growth factor, could not overcome the inhibitory action of PAF in wound closure. Three PAF antagonists, BN50727, BN50730, and BN50739, abolished the effect of PAF. A significant increase in TUNEL-positive staining occurred in corneal stromal cells (keratocytes), which was inhibited by preincubating the corneas with PAF antagonists. However, no TUNEL-positive staining was found in epithelial cells. TUNEL-staining results in cultured stromal cells (keratocytes) and epithelial cells in first-passage cell culture were similar to those in organ-cultured corneas. In addition, PAF caused 35% to 56% inhibition of adhesion of epithelial cells to proteins of the extracellular matrix: collagen I and IV, fibronectin, and laminin. There were no significant changes in proliferation or migration of epithelial cells induced by the lipid mediator. The results suggest PAF plays an important role in preventing corneal wound healing by affecting adhesion of epithelial cells and increasing apoptosis in stromal cells. PAF antagonists could be of therapeutic importance during prolonged ocular inflammation, helping to avoid loss of corneal transparency and visual acuity.

Human Asymptomatic Epitope Peptide/CXCL10-Based Prime/Pull Vaccine Induces Herpes Simplex Virus-Specific Gamma Interferon-Positive CD107+ CD8+ T Cells That Infiltrate the Cornea and Trigeminal Ganglia of Humanized HLA Transgenic Rabbits and Protect against Ocular Herpes Challenge.

PubMed

Khan, Arif A; Srivastava, Ruchi; Vahed, Hawa; Roy, Soumyabrata; Walia, Sager S; Kim, Grace J; Fouladi, Mona A; Yamada, Taikun; Ly, Vincent T; Lam, Cynthia; Lou, Anthony; Nguyen, Vivianna; Boldbaatar, Undariya; Geertsema, Roger; Fraser, Nigel W; BenMohamed, Lbachir

2018-06-13

Herpes simplex virus 1 (HSV-1) is a prevalent human pathogen that infects the cornea causing potentially blinding herpetic disease. A clinical herpes vaccine is still lacking. In the present study, a novel prime/pull vaccine was tested in Human Leukocyte Antigen- (HLA-) transgenic rabbit model of ocular herpes (HLA Tg rabbit). Three asymptomatic (ASYMP) peptide epitopes were selected from the HSV-1 membrane glycoprotein C (UL44 400-408 ), the DNA replication binding helicase (UL9 196-204 ), and the tegument protein (UL25 572-580 ), all preferentially recognized by CD8 + T cells from "naturally protected" HSV-1-seropositive healthy ASYMP individuals (who never had recurrent corneal herpetic disease). HLA Tg rabbits were immunized with a mixture of these three ASYMP CD8 + T cell peptide epitopes (UL44 400-408 , UL9 196-204 and UL25 572-580 ), delivered subcutaneously with CpG 2007 adjuvant (prime). Fifteen days later, half of the rabbits received a topical ocular treatment with a recombinant neurotropic AAV8 vector, expressing the T cell-attracting CXCL10 chemokine (pull). The frequency, function of HSV-specific CD8 + T cells induced by the prime/pull vaccine were assessed in peripheral blood, cornea, and trigeminal ganglia (TG). Compared to peptides alone, the peptides/CXCL10 prime/pull vaccine generated frequent polyfunctional gamma interferon-positive (IFN-γ + ) CD107 + CD8 + T cells that infiltrated both the cornea and TG. CD8 + T cells mobilization into cornea and TG of prime/pull- vaccinated rabbits was associated with a significant reduction in corneal herpes infection and disease following an ocular HSV-1 challenge (McKrae). These findings draw attention to the novel prime/pull vaccine strategy to mobilize anti-viral CD8 + T cells into tissues protecting them against herpes infection and disease. IMPORTANCE There is an urgent need for a vaccine against widespread herpes simplex virus infections. The present study demonstrates that immunization of HLA transgenic rabbits with a peptide/CXCL10 prime/pull vaccine triggered mobilization of HSV-specific CD8 + T cells locally in the cornea and TG, the sites of acute and latent herpes infections. Mobilization of antiviral CD8 + T cells into cornea and TG of rabbits that received the prime/pull vaccine was associated with protection against ocular herpes infection and disease following an ocular HSV-1 challenge. These results highlight the importance of the prime/pull vaccine strategy to bolster the number and function of protective CD8 + T cells within infected tissues. Copyright © 2018 American Society for Microbiology.

Imaging shear stress distribution and evaluating the stress concentration factor of the human eye

NASA Astrophysics Data System (ADS)

Joseph Antony, S.

2015-03-01

Healthy eyes are vital for a better quality of human life. Historically, for man-made materials, scientists and engineers use stress concentration factors to characterise the effects of structural non-homogeneities on their mechanical strength. However, such information is scarce for the human eye. Here we present the shear stress distribution profiles of a healthy human cornea surface in vivo using photo-stress analysis tomography, which is a non-intrusive and non-X-ray based method. The corneal birefringent retardation measured here is comparable to that of previous studies. Using this, we derive eye stress concentration factors and the directional alignment of major principal stress on the surface of the cornea. Similar to thermometers being used for monitoring the general health in humans, this report provides a foundation to characterise the shear stress carrying capacity of the cornea, and a potential bench mark for validating theoretical modelling of stresses in the human eye in future.

In vivo laser confocal microscopic analysis of corneal K-structures after keratorefractive surgery (LASIK and epi-LASIK).

PubMed

Yokogawa, Hideaki; Kobayashi, Akira; Tagawa, Kosaku; Sugiyama, Kazuhisa

2010-01-01

To demonstrate alterations of corneal K-structures (sub-Bowman's fibrous structures) after keratorefractive surgery by in vivo laser confocal microscopy and to look for association of K-structures with fluorescein-stained anterior corneal mosaic (ACM). Five patients (nine eyes) participated in this study. For four patients, one eye was evaluated after laser in situ keratomileusis (LASIK) and the other after epipolis-laser in situ keratomileusis (epi-LASIK). For one patient, the left eye was evaluated after epithelial debridement. A photograph of the ACM was obtained. Central corneal regions were scanned by Heidelberg Retina Tomograph 2 Rostock Cornea Module (Heidelberg Engineering GmbH, Heidelberg, Germany). The ACM and K-structures disappeared in all corneas after epi-LASIK, but not after LASIK and epithelial debridement cornea. The presence of K-structures and ACM may be an index to identify eyes that had a previous refractive surgical procedure (surface ablation or LASIK) and be a health index of Bowman layer and adjacent anterior stroma. Copyright 2010, SLACK Incorporated.

THz and mm-Wave Sensing of Corneal Tissue Water Content: In Vivo Sensing and Imaging Results

PubMed Central

Taylor, Zachary D.; Garritano, James; Sung, Shijun; Bajwa, Neha; Bennett, David B.; Nowroozi, Bryan; Tewari, Priyamvada; Sayre, James W.; Hubschman, Jean-Pierre; Deng, Sophie X.; Brown, Elliott R.; Grundfest, Warren S.

2015-01-01

A pulsed terahertz (THz) imaging system and millimeter-wave reflectometer were used to acquire images and point measurements, respectively, of five rabbit cornea in vivo. These imaging results are the first ever produced of in vivo cornea. A modified version of a standard protocol using a gentle stream of air and a Mylar window was employed to slightly dehydrate healthy cornea. The sensor data and companion central corneal thickness (CCT) measurements were acquired every 10–15 min over the course of two hours using ultrasound pachymmetry.. Statistically significant positive correlations were established between CCT measurements and millimeter wave reflectivity. Local shifts in reflectivity contrast were observed in the THz imagery; however, the THz reflectivity did not display a significant correlation with thickness in the region probed by the 100 GHz and CCT measurements. This is explained in part by a thickness sensitivity at least 10× higher in the mm-wave than the THz systems. Stratified media and effective media modeling suggest that the protocol perturbed the thickness and not the corneal tissue water content (CTWC). To further explore possible etalon effects, an additional rabbit was euthanized and millimeter wave measurements were obtained during death induced edema. These observations represent the first time that the uncoupled sensing of CTWC and CCT have been achieved in vivo. PMID:26161292

Internal and external contamination of donor corneas before in situ excision: bacterial risk factors in 93 donors.

PubMed

Robert, Pierre-Yves; Camezind, Philippe; Drouet, Mireille; Ploy, Marie-Cécile; Adenis, Jean-Paul

2002-04-01

Most studies of corneal donor contamination concentrate on postenucleation contamination of the eyeball. The purpose of the present study was to evaluate the relative contamination of in situ excised corneal tissue and relevance to final success or rejection by recipients of the corneal grafts. Ninety-three donors underwent anterior chamber puncture (ACP) and corneal epithelium scarification (CS) before and after disinfection with 5% povidone-iodine. Following in situ excision, corneas were preserved in culture medium at +35 degrees C. Morphological and bacteriological assessment was carried out after culture, and recipients were followed up over a 2-year period. Samples taken by ACP, CS before disinfection, CS after disinfection and a culture medium sample were contaminated by bacteria in, respectively, 8 (8.6%), 23 (24.7%) 4 (4.3%) and 5 (5.4%) donors. Contamination of aqueous humor was significantly associated with age, death-to-sample time and premortem systemic infection. Contamination of epithelium significantly increased culture medium contamination. External bacteria on donor cornea are mainly skin bacteria (especially Staphylococcus) and can be partially eliminated by a povidone-iodine wash. Internal bacteria are mainly gut bacteria and may be due to perimortem bacteriemia. However, bacterial infection at the time of death appears to have no effect on the incidence of endophthalmitis in recipients and should no longer prevent use of such corneal tissue in grafts.

[Costs of preserved corneal transplants].

PubMed

Ardjomand, N; Reich, M E

1997-10-01

Organ culture medium and Optisol are the most commonly used corneal storage mediums. This study compares the costs for these two methods. In the calculation of costs we did not just take the direct costs into account, but also tried to determine the fixed costs per transplanted cornea with corresponding assumptions. Proceeding on the assumption that 50 stored corneas were transplanted per year, an amount of 11,660 ATS (1,666 DM, 857 ECU) for each organ cultured and 11,986 ATS (1,712 DM, 881 ECU) for each graft preserved in Optisol was calculated. Raising the number of transplanted corneas to 400 per year, each tissue stored in organ culture medium costs 2,811 ATS (402 DM, 207 ECU) and those preserved in Optisol 3234 ATS (462 DM, 238 ECU). Since organ culture storage gives us a reduction in costs of more than 15% compared to storing in Optisol, when preserving 400 transplantable grafts, from the business economics aspect, this storage method should be preferred.

The G-protein-coupled receptor, GPR84, is important for eye development in Xenopus laevis.

PubMed

Perry, Kimberly J; Johnson, Verity R; Malloch, Erica L; Fukui, Lisa; Wever, Jason; Thomas, Alvin G; Hamilton, Paul W; Henry, Jonathan J

2010-11-01

G-protein-coupled receptors (GPCRs) represent diverse, multifamily groups of cell signaling receptors involved in many cellular processes. We identified Xenopus laevis GPR84 as a member of the A18 subfamily of GPCRs. During development, GPR84 is detected in the embryonic lens placode, differentiating lens fiber cells, retina, and cornea. Anti-sense morpholino oligonucleotide-mediated knockdown and RNA rescue experiments demonstrate GPR84's importance in lens, cornea, and retinal development. Examination of cell proliferation using an antibody against histone H3 S10P reveals significant increases in the lens and retina following GPR84 knockdown. Additionally, there was also an increase in apoptosis in the retina and lens, as revealed by TUNEL assay. Reciprocal transplantation of the presumptive lens ectoderm between uninjected controls and morpholino-injected embryos demonstrates that GPR84 is necessary in the retina for proper development of the retina, as well as other eye tissues including the lens and cornea. © 2010 Wiley-Liss, Inc.

The G-protein-coupled receptor, GPR84, is important for eye development in Xenopus laevis

PubMed Central

Perry, Kimberly J.; Johnson, Verity R.; Malloch, Erica L.; Fukui, Lisa; Wever, Jason; Thomas, Alvin G.; Hamilton, Paul W.; Henry, Jonathan J.

2010-01-01

G-protein-coupled receptors (GPCRs) represent diverse, multifamily groups of cell signaling receptors involved in many cellular processes. We identified Xenopus laevis GPR84 as a member of the A18 subfamily of GPCRs. During development, GPR84 is detected in the embryonic lens placode, differentiating lens fiber cells, retina and cornea. Anti-sense morpholino oligonucleotide-mediated knockdown and RNA rescue experiments demonstrate GPR84’s importance in lens, cornea and retinal development. Examination of cell proliferation using an antibody against histone H3 S10P reveals significant increases in the lens and retina following GPR84 knockdown. Additionally, there was also an increase in apoptosis in the retina and lens, as revealed by TUNEL assay. Reciprocal transplantation of the presumptive lens ectoderm between uninjected controls and morpholino injected embryos demonstrates that GPR84 is necessary in the retina for proper development of the retina, as well as other eye tissues including the lens and cornea. PMID:20925114

T-style keratoprosthesis based on surface-modified poly (2-hydroxyethyl methacrylate) hydrogel for cornea repairs.

PubMed

Xiang, Jun; Sun, Jianguo; Hong, Jiaxu; Wang, Wentao; Wei, Anji; Le, Qihua; Xu, Jianjiang

2015-05-01

Corneal disease is a common cause of blindness, and keratoplasty is considered as an effective treatment method. However, there is a severe shortage of donor corneas worldwide. This paper presents a novel T-style design of a keratoprosthesis and its preparation methods, in which a mechanically and structurally effective artificial cornea is made based on a poly(2-hydroxyethyl methacrylate) hydrogel. The porous skirt was modified with hyaluronic acid and cationized gelatin, and the bottom of the optical column was coated with poly(ethylene glycol). The physical properties of the T-style Kpro were analyzed using ultraviolet and visible spectrophotometry and electron scanning microscopy. The surface chemical properties were characterized using Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy. The surface modification in the spongy skirt promoted cell adhesion and produced a firm bond between the corneal tissue and the implant device, while the surface modification in the optic column resisted cell adhesion and prevented retroprosthetic membrane formation. Through improved surgical techniques, the novel T-style keratoprosthesis provides enough mechanical stability to facilitate long-term biointegration with the host environment. In vivo implantation experiments showed that the T-style keratoprosthesis is a promising cornea alternative for patients with severe limbal stem cell deficiency and corneal opacity. Copyright © 2015 Elsevier B.V. All rights reserved.

Corneal elastic anisotropy and hysteresis as a function of IOP assessed by optical coherence elastography

NASA Astrophysics Data System (ADS)

Li, Jiasong; Singh, Manmohan; Han, Zhaolong; Wu, Chen; Raghunathan, Raksha; Liu, Chih-Hao; Nair, Achuth; Noorani, Shezaan; Aglyamov, Salavat R.; Twa, Michael D.; Larin, Kirill V.

2016-03-01

The mechanical anisotropic properties of the cornea can be an important indicator for determining the onset and severity of different diseases and can be used to assess the efficacy of various therapeutic interventions, such as cross-linking and LASIK surgery. In this work, we introduce a noncontact method of assessing corneal mechanical anisotropy as a function of intraocular pressure (IOP) using optical coherence elastography (OCE). A focused air-pulse induced low amplitude (<10 μm) elastic waves in fresh porcine corneas in the whole eye-globe configuration in situ. A phase-stabilized swept source optical coherence elastography (PhS-SSOCE) system imaged the elastic wave propagation at stepped radial angles, and the OCE measurements were repeated as the IOP was cycled. The elastic wave velocity was then quantified to determine the mechanical anisotropy and hysteresis of the cornea. The results show that the elastic anisotropy at the corneal of the apex of the cornea becomes more pronounced at higher IOPs, and that there are distinct radial angles of higher and lower stiffness. Due to the noncontact nature and small amplitude of the elastic wave, this method may be useful for characterizing the elastic anisotropy of ocular and other tissues in vivo completely noninvasively.

Study of transmittance and reflectance spectra of the cornea and the sclera in the THz frequency range

NASA Astrophysics Data System (ADS)

Iomdina, Elena N.; Goltsman, Gregory N.; Seliverstov, Sergey V.; Sianosyan, Alisa A.; Teplyakova, Kseniya O.; Rusova, Anastasia A.

2016-09-01

An adequate water balance (hydration extent) is one of the basic factors of normal eye function, including its external shells: the cornea and the sclera. Adequate control of corneal and scleral hydration is very important for early diagnosis of a variety of eye diseases, stating indications for and contraindications against keratorefractive surgeries and the choice of contact lens correction solutions. THz systems of creating images in reflected beams are likely to become ideal instruments of noninvasive control of corneal and scleral hydration degrees. This paper reports on the results of a study involving transmittance and reflectance spectra for the cornea and the sclera of rabbit and human eyes, as well as those of the rabbit eye, in the frequency range of 0.13 to 0.32 THz. The dependence of the reflectance coefficient of these tissues on water mass percentage content was determined. The experiments were performed on three corneas, three rabbit scleras, two rabbit eyes, and three human scleras. The preliminary results demonstrate that the proposed technique, based on the use of a continuous THz radiation, may be utilized to create a device for noninvasive control of corneal and scleral hydration, which has clear potential of broad practical application.

Transdifferentiation from cornea to lens in Xenopus laevis depends on BMP signalling and involves upregulation of Wnt signalling

PubMed Central

2011-01-01

Background Surgical removal of the lens from larval Xenopus laevis results in a rapid transdifferention of central corneal cells to form a new lens. The trigger for this process is understood to be an induction event arising from the unprecedented exposure of the cornea to the vitreous humour that occurs following lens removal. The molecular identity of this trigger is unknown. Results Here, we have used a functional transgenic approach to show that BMP signalling is required for lens regeneration and a microarray approach to identify genes that are upregulated specifically during this process. Analysis of the array data strongly implicates Wnt signalling and the Pitx family of transcription factors in the process of cornea to lens transdifferentiation. Our analysis also captured several genes associated with congenital cataract in humans. Pluripotency genes, in contrast, were not upregulated, supporting the idea that corneal cells transdifferentiate without returning to a stem cell state. Several genes from the array were expressed in the forming lens during embryogenesis. One of these, Nipsnap1, is a known direct target of BMP signalling. Conclusions Our results strongly implicate the developmental Wnt and BMP signalling pathways in the process of cornea to lens transdifferentiation (CLT) in Xenopus, and suggest direct transdifferentiation between these two anterior eye tissues. PMID:21896182

Toxicological effects and recovery of the corneal epithelium in Cyprinus carpio communis Linn. exposed to monocrotophos: an scanning electron microscope study.

PubMed

Uppal, Ravneet Kaur; Johal, Mohinder Singh; Sharma, Madan Lal

2015-05-01

This study was conducted based on the evidence of fish habitats in North India being affected by organophosphate pesticides draining from agricultural fields into bodies of water, especially during the rainy season. Various tissues of fish such as scales, gills ovaries, kidney, and liver have been studied from the toxicological point of view, but the toxicological effects of aquatic pollutants on fish cornea have not been investigated to date. We conducted comparative toxicological studies on the cornea of Cyprinus carpio communis using two sublethal (0.038 and 0.126 ppm) concentrations of monocrotophos pesticide for 30 days. Corneas from all the groups were evaluated by a scanning electron microscope. The fish exposed to the monocrotophos pesticide developed corneal necrosis due to the formation of crystalloid-like structures, thinning and shrinkage of microridges on the corneal epithelium. After 30 days, fish from the monocrotophos-treated tank were transferred to normal environmental conditions. After 60 days under natural condition, epithelial cells did not fully recover. In conclusion, exposure to monocrotophos induces irreversible changes in the cornea of C. carpio communis. As fish and mammalian visual systems share many similarities, the reported finding may offer useful insights for further toxicological and ophthalmological studies in humans. © 2013 American College of Veterinary Ophthalmologists.

Ultraviolet A: Visible spectral absorbance of the human cornea after transepithelial soaking with dextran-enriched and dextran-free riboflavin 0.1% ophthalmic solutions.

PubMed

Lombardo, Marco; Micali, Norberto; Villari, Valentina; Serrao, Sebastiano; Pucci, Giuseppe; Barberi, Riccardo; Lombardo, Giuseppe

2015-10-01

To evaluate the stromal concentration of 2 commercially available transepithelial riboflavin 0.1% solutions in human donor corneas with the use of spectrophotometry. University of Calabria, Rende, Italy. Experimental study. The absorbance spectra of 12 corneal tissues were measured in the 330 to 700 nm wavelength range using a purpose-designed spectrophotometry setup before and after transepithelial corneal soaking with a 15% dextran-enriched riboflavin 0.1% solution (n = 6) or a hypotonic dextran-free riboflavin 0.1% solution (n = 6). Both ophthalmic solutions contained ethylenediaminetetraacetic acid and trometamol as enhancers. In addition, 4 deepithelialized corneal tissues underwent stromal soaking with a 20% dextran-enriched riboflavin 0.1% solution and were used as controls. All the riboflavin solutions were applied topically for 30 minutes. The stromal concentration of riboflavin was quantified by analysis of absorbance spectra of the cornea collected before and after application of each solution. The mean stromal riboflavin concentration was 0.012% ± 0.003% (SD), 0.0005% ± 0.0003% (P < .001), and 0.004% ± 0.001% (P < .01) in tissues soaked with 20% dextran-enriched, 15% dextran-enriched, and hypotonic dextran-free solutions, respectively. The difference of stromal riboflavin concentration between the 2 transepithelial solutions was statistically significant (P < .01). Dextran-enriched solutions required complete corneal deepithelialization to permit effective stromal soaking with riboflavin. Nevertheless, riboflavin in hypotonic dextran-free solution with enhancers permeates across stroma through an intact epithelium. No author has a financial or proprietary interest in any material or method mentioned. Copyright © 2015 ASCRS and ESCRS. Published by Elsevier Inc. All rights reserved.

Ocular and systemic pharmacokinetics of lidocaine hydrochloride ophthalmic gel in rabbits after topical ocular administration.

PubMed

Liu, Bing; Ding, Li; Xu, Xiaowen; Lin, Hongda; Sun, Chenglong; You, Linjun

2015-12-01

Lidocaine hydrochloride ophthalmic gel is a novel ophthalmic preparation for topical ocular anesthesia. The study is aimed at evaluating the ocular and systemic pharmacokinetics of lidocaine hydrochloride 3.5 % ophthalmic gel in rabbits after ocular topical administration. Thirty-six rabbits were randomly placed in 12 groups (3 rabbits per group). The rabbits were quickly killed according to their groups at 0 (predose), 0.0833, 0.167, 0.333, 0.667, 1, 1.5, 2, 3, 4, 6, and 8 h postdose and then the ocular tissue and plasma samples were collected. All the samples were analyzed by a validated LC-MS/MS method. The test result showed that the maximum concentration (C max) of lidocaine in different ocular tissues and plasma were all achieved within 20 min after drug administration, and the data of C max were (2,987 ± 1814) μg/g, (44.67 ± 12.91) μg/g, (26.26 ± 7.19) μg/g, (11,046 ± 2,734) ng/mL, and (160.3 ± 61.0) ng/mL for tear fluid, cornea, conjunctiva, aqueous humor, and plasma, respectively. The data of the elimination half-life in these tissues were 1.5, 3.2, 3.5, 1.9, and 1.7 h for tear fluid, cornea, conjunctiva, aqueous humor, and plasma, respectively. The intraocular lidocaine levels were significantly higher than that in plasma, and the elimination half-life of lidocaine in cornea, conjunctiva, and aqueous humor was relatively longer than that in tear fluid and plasma. The high intraocular penetration, low systemic exposure, and long duration in the ocular tissues suggested lidocaine hydrochloride 3.5 % ophthalmic gel as an effective local anesthetic for ocular anesthesia during ophthalmic procedures.

Dextran Preserves Native Corneal Structure During Decellularization.

PubMed

Lynch, Amy P; Wilson, Samantha L; Ahearne, Mark

2016-06-01

Corneal decellularization has become an increasingly popular technique for generating scaffolds for corneal regeneration. Most decellularization procedures result in tissue swelling, thus limiting their application. Here, the use of a polysaccharide, dextran, to reduce swelling and conserve the native corneal structure during decellularization was investigated. Corneas were treated with 1% Triton X-100, 0.5% sodium dodecyl sulfate, and nucleases under constant rotation followed by extensive washing. To reduce swelling, decellularization solutions were supplemented with 5% dextran either throughout the whole decellularization process or during the washing cycles only. Quantitative analysis of DNA content showed a 96% reduction after decellularization regardless of the addition of dextran. Dextran resulted in a significant reduction in swelling from 3.85 ± 0.43 nm without to 1.94 ± 0.29-2.01 ± 0.37 nm (p < 0.05) remaining at similar dimensions to the native tissue (1.73 ± 0.23 nm). Tissue transparency was restored to all decellularized corneas following submersion in glycerol. Transmission electron microscopy (TEM) analysis found that dextran must be present throughout the decellularization protocol to preserve the native corneal architecture, anisotropy analysis demonstrated comparable results (0.22 ± 0.03) to the native cornea (0.24 ± 0.02), p > 0.05. Dextran can counteract the detrimental effects of decellularizing agents on the biomechanical properties of the tissue resulting in similar compressive moduli (mean before decellularization: 5.40 ± 1.18 kPa; mean after decellularization with dextran: 5.64 ± 1.34 kPa, p > 0.05). Cells remained viable in the presence of decellularized scaffolds. The findings of this study indicate that dextran not only prevents significant corneal swelling during decellularization but also enhances the maintenance of the native corneal ultrastructure.

Single-Pass Microkeratome System for Eye Bank DSAEK Tissue Preparation: Is Stromal Bed Thickness Predictable and Reproducible?

PubMed

Choulakian, Mazen Y; Li, Jennifer Y; Ramos, Samuel; Mannis, Mark J

2016-01-01

To evaluate the predictability and reproducibility of stromal bed thickness for single-pass donor Descemet stripping automated endothelial keratoplasty (DSAEK) tissue preparation, using the ML7 Microkeratome Donor Cornea System (Med-logics Inc, Athens, TX). In this retrospective chart review of 256 consecutive corneal tissue preparations for DSAEK surgery, from June 2013 to August 2014, tissue thicknesses were divided into 3 groups, depending on surgeon preference: <91 μm (group A), 90 to 120 μm (group B), and 120 to 160 μm (group C). Precut and postcut data were recorded. Average postcut donor corneal thickness was 114 ± 30 μm (range 60-183 μm), whereas the average in group A was 97 ± 23 μm (range 60-128), in group B was 113 ± 21 μm (range 77-179), and in group C was 134 ± 43 (range 89-183). Average postcut endothelial cell density was very adequate at 3013 ± 250 cells per square millimeter. There were a total of 7 failed procedures from 256 attempts, which represents a rate of 2.7%. This rate decreases to 1.5% when analyzing the last 200 cuts. The ML7 Microkeratome Donor Cornea System allows for reliable and reproducible DSAEK tissue preparation. Ultrathin DSAEK tissues can be prepared with a single-pass. Aiming for a graft thickness between 90 and 120 μm seems to be most reliable.

Donation transplants and tissue banking in Mexico.

PubMed

Luna-Zaragoza, D; Reyes-Frías, M L

2001-01-01

Knowledge about transplants in Mexico goes back to the Aztec period. Today the need for organ and tissue transplants in Mexico is high; the estimated number is 100,000 patients, but there are only 2 donors per million population, for corneas. The organ, tissue and cell transplantation law which was modified in 2,000, establishes that when a person dies, he will be a potential donor of organs and tissues. This new law will give hope to many patients, since it is expected to increase significantly the amount of organs and tissues for transplants. At present Mexico has 178 hospitals that are authorized to carry out organ and tissue transplants, and 53 Tissue Banks.

The Resistance of Certain Tissues to Invasion

PubMed Central

Eisenstein, Reuben; Sorgente, Nino; Soble, Lawrence W.; Miller, Alexander; Kuettner, Klaus E.

1973-01-01

If puppy tissues are explanted onto the chick chorioallantoic membrane, those tissues which normally have a blood supply are rapidly invaded by vascularized mesenchyme of host origin. Hyaline cartilage, a tissue virtually devoid of blood vessels, is impenetrable by proliferating mesenchyme of the host, while calcified cartilage, which normally is vascularized, is penetrable. The stroma of the cornea, another normally avascular tissue, is readily penetrable, but Descemet's membrane forms a barrier to invasion by host tissues. The experimental system used permits the design of experiments in which the study of factors responsible for the resistance of tissues such as cartilage to invasion can be undertaken. ImagesFig 1Fig 2Fig 3Fig 4 PMID:4129060

Intergrin-dependent neutrophil migration in the injured mouse cornea

USDA-ARS?s Scientific Manuscript database

As an early responder to an inflammatory stimulus, neutrophils must exit the vasculature and migrate through the extravascular tissue to the site of insult, which is often remote from the point of extravasation. Following a central epithelial corneal abrasion, neutrophils recruited from the peripher...

The surface of the eye--a superficial entity with deep repercussions.

PubMed

Potop, Vasile; Dumitrache, Marieta; Ciocalteu, Alina

2009-01-01

The surface of the eye is an anatomical and functional entity with a relatively recent delimitation but with significant therapeutic and diagnostic consequences. The pathology of the conjunctive and cornea must be approached by looking at the interrelations between the two tissues that are so different anatomically and functionally but in the same time form a unit in structuring the eye's surface. There are two major categories of relations between the two tissues: one of them is mediated by lachrymal secretion, a process whose complexity is not yet fully understood, and the other is germinal, referring to the stem cells located at the limbus which become epithelial cornea cells that can fixate lachrymal fluid. Imbalances in the quantity and quality of lachrymal secretion can be compensated, up to a certain point, by artificial products, but in severe cases only specially prepared autologous serum can compensate the deficit. The limbic deficits that affect stem cells require complex therapeutic procedures like limbic cell transplant, using an amniotic membrane or autologous serum.

Iron homeostasis and eye disease

PubMed Central

Loh, Allison; Hadziahmetovic, Majda; Dunaief, Joshua L.

2009-01-01

Summary Iron is necessary for life, but excess iron can be toxic to tissues. Iron is thought to damage tissues primarily by generating oxygen free radicals through the Fenton reaction. We present an overview of the evidence supporting iron's potential contribution to a broad range of eye disease using an anatomical approach. Firstly, iron can be visualized in the cornea as iron lines in the normal aging cornea as well as in diseases like keratoconus and pterygium. In the lens, we present the evidence for the role of oxidative damage in cataractogenesis. Also, we review the evidence that iron may play a role in the pathogenesis of the retinal disease age-related macular degeneration. Although currently there is no direct link between excess iron and development of optic neuropathies, ferrous iron's ability to form highly reactive oxygen species may play a role in optic nerve pathology. Lastly, we discuss recent advances in prevention and therapeutics for eye disease with antioxidants and iron chelators,. PMID:19059309

Ultraviolet laser effects on the cornea

NASA Astrophysics Data System (ADS)

Zuclich, Joseph A.

1990-07-01

Ultraviolet radiation in the ambient environment or from artificial sources may pose both acute and chronic hazards to the skin and the ocular tissues. In general terrestrial conditions have evolved such that there are only narrow safety margins between ambient UV levels and exposure levels harmful to the human. Obvious examples of acute consequences ofUV overexposure are sunburn and snowblindness as well as analogous conditions induced by artificial sources such as the welder''s arc mercury vapor lamps and UV-emitting lasers. Further chronic UV exposure is strongly implicated as a causative agent in certain types of cataract and skin cancer. This presentation will summarize a number of specific cases where UV radiation affected the primate cornea. Data presented will include the action spectra for far- and near-UV induced ocular damage the pulsewidth and total energy dependencies of ocular thresholds studies of cumulative effects of repeated UV exposures and quantitative determinations of tissue repair or recovery rates. Depending on the exposure parameters utilized photochemical thermal or photoablative damage mechanisms may prevail. 1.

Collagen a natural scaffold for biology and engineering

USDA-ARS?s Scientific Manuscript database

Collagen, the most abundant protein in mammals, constitutes a quarter of the animal's total weight. The unique structure of fibrous collagens, a long triple helix that further associates into fibers, provides an insoluble scaffold that gives strength and form to the skin, tendons, bones, cornea and...

The cornea in connective tissue diseases.

PubMed

Maumenee, I H

1978-10-01

Even though lenticular and retinal abnormalities seem to dominate the ophthalmologic picture in Marfan's syndrome, the cornea shows significant abnormalities consisting of a striking flattening and corneal astigmatism. The use of conjunctival biopsies followed by histochemical and electron-microscopic evaluation shows low morbidity but an excellent yield of diagnostic information on storage diseases, and an ectopic collagen may be the basis of at least one type of keratoconus. Much more work has to be done on defining the collagens of the human eye in embryologic, fetal, and postnatal stages under normal and pathologic conditions. The yield of such studies may be high for an understanding of such diseases as myopia, retinal detachment, and keratoconus.

Ultrastructure and mineral composition of the cornea cuticle in the compound eyes of a supralittoral and a marine isopod.

PubMed

Alagboso, Francisca I; Reisecker, Christian; Hild, Sabine; Ziegler, Andreas

2014-08-01

The cuticle of the cornea in Crustacea is an interesting example of a composite material compromising between two distinct functions. As part of the dioptric apparatus of the ommatidia within the complex eye it forms transparent micro-lenses that should as well maintain the mechanical stability of the head capsule. We analyzed the ultrastructure and composition of the isopod cornea cuticle of the terrestrial species Ligia oceanica and the marine Sphaeroma serratum. We used a variety of tissue preparation methods, electron microscopic techniques as well as electron microprobe analysis and Raman spectroscopic imaging. The results reveal various structural adaptations that likely increase light transmission. These are an increase in the thickness of the epicuticle, a reduction of the thickness of the outer layer of calcite, a spatial restriction of pore canals to interommatidial regions, and, for S. serratum only, an increase in calcite crystal size. In both species protein-chitin fibrils within the proximal exocuticle form a peculiar reticular structure that does not occur within the cuticle of the head capsule. In L. oceanica differential mineralization results in a spherically shaped interface between mineralized and unmineralized endocuticle, likely an adaptation to increase the refractive power of the cornea maintaining the mechanical stability of the cuticle between the ommatidia. The results show that the habitat and differences in the general structure of the animal's cuticle affect the way in which the cornea is adapted to its optical function. Copyright © 2014 Elsevier Inc. All rights reserved.

Long-term consequences of topical dexamethasone treatment during acute corneal HSV-1 infection on the immune system

PubMed Central

Chucair-Elliott, Ana J.; Carr, Meghan M.; Carr, Daniel J. J.

2017-01-01

Herpes simplex virus type 1 (HSV-1) is a leading cause of neurotrophic keratitis (NTK). NTK is characterized by decreased corneal sensation from damage to the corneal sensory fibers. We have reported on the regression of corneal nerves and their function during acute HSV-1 infection. That nerve loss is followed by an aberrant process of nerve regeneration during the latent phase of infection that lacks functional recovery. We recently showed the elicited immune response in the infected cornea, and not viral replication itself, is part of the mechanism responsible for the nerve degeneration process after infection. Specifically, we showed infected corneas topically treated with dexamethasone (DEX) significantly retained both structure and sensitivity of the corneal nerve network in comparison to mice treated with control eye drops, consistent with decreased levels of proinflammatory cytokines and reduced influx of macrophages and CD8+ T cells into the cornea. This study was undertaken to analyze the long-term effect of such a localized, immunosuppressive paradigm (DEX drops on the cornea surface during the first 8 d of HSV-1 infection) on the immune system and on corneal pathology. We found the profound immunosuppressive effect of DEX on lymphoid tissue was sustained in surviving mice for up to 30 d postinfection (p.i.). DEX treatment had prolonged effects, preserving corneal innervation and its function and blunting neovascularization, as analyzed at 30 d p.i. Our data support previously reported observations of an association between the persistent presence of inflammatory components in the latently infected cornea and structural and functional nerve defects in NTK. PMID:28115476

Assessing the Impact of Diabetes Mellitus on Donor Corneal Endothelial Cell Density.

PubMed

Liaboe, Chase A; Aldrich, Benjamin T; Carter, Pamela C; Skeie, Jessica M; Burckart, Kimberlee A; Schmidt, Gregory A; Reed, Cynthia R; Zimmerman, M Bridget; Greiner, Mark A

2017-05-01

To quantify changes in endothelial cell density (ECD) of donor corneal tissue in relation to the presence or absence of a medical history of diabetes mellitus diagnosis, treatment, and complications. A retrospective review was performed for all corneas collected at Iowa Lions Eye Bank between January 2012 and December 2015. For purposes of analysis, donor corneas were divided into 4 groups: nondiabetic, non-insulin-dependent diabetic, insulin-dependent diabetic without medical complications due to diabetes, and insulin-dependent diabetic with medical complications due to diabetes. ECD values (obtained through specular microscopy) and transplant suitability for endothelial transplantation (determined by the standard protocol of the eye bank) were compared among groups using linear mixed model analysis. In total, 4185 corneas from 2112 donors were included for analysis. Insulin-dependent diabetic samples with medical complications due to diabetes (N = 231 from 119 donors) showed lower ECD values compared with nondiabetic samples (-102 cells/mm, P = 0.049) and non-insulin-dependent diabetic samples (-117 cells/mm, P = 0.031). ECD values did not differ significantly among the remaining groups. The likelihood of suitability for endothelial transplantation did not differ among all 4 groups. Corneas from donors with insulin-dependent diabetes mellitus and medical complications resulting from the disease have lower mean ECD values compared with other donors. However, our analysis suggests that these corneas are equally likely to be included in the donor pool for corneal transplantation. Additional studies are needed to determine the mechanism(s) contributing to cell loss in donors with advanced diabetes and to assess associated endothelial cell functional impairment.

Specific demographic factors could predict deceased potential cornea donors: A retrospective study from Beijing Tongren Hospital Eye Bank.

PubMed

Liu, Yang; Zhang, Yingnan; Zhang, Jing; Yu, Jin; Liang, Qingfeng; Pan, Zhiqiang

2017-12-01

Compared with evident cornea donors (ECDs), deceased potential cornea donors (DPCDs) have no obvious donor identifications to reference, which causes many eligible cornea tissues to be wasted. The demographic characteristics of DPCDs might be different from those of ECDs owing to the following different features: donation consent provided by relatives and willingness to donate before death. Thus, the aim of this study is to reveal the demographic characteristics of DPCDs by comparing DPCDs and ECDs.The demographic factors of 138 donors (both DPCDs and ECDs) were collected from the Beijing Tongren Hospital Eye Bank database and analyzed. To differentiate DPCDs from ECDs using the above-mentioned features, we interviewed the relatives of the donors by telephone. The relatives' attitudes toward cornea donation and their suggestions for our donation service were also acquired during the interview. Two logistic regressions were performed to reveal the demographic factors influencing the 2 features and indicate DPCDs.The donors had certain demographic characteristics (elderly, secondary, or tertiary education level, central district resident), and the most frequent cause of death for the donors was a malignant tumor (n = 56, 43.1%). All the relatives had positive attitudes toward cornea donations, and they hoped to increase publicity efforts to encourage more people to donate and establish more convenient and efficient access for cornea donation. In univariate regressions, age (P = .004, >50 years: odds ratio [OR] = 6.89, 95% confidence interval [CI]: 1.82-26.05), marital status (P = .043, divorced: OR = 9.00,95% CI: 1.33-60.80) significantly influenced relative consent, whereas age (P = .001, >50 years: OR = 15.00, 95% CI: 3.00-74.98), and family address (P = .001, central district: OR = 1) were significant factors influencing the willingness to donate before death. In multivariate regression, age (P = .021, >50 years: OR = 8.14, 95% CI: 1.37-48.41) was the only significant factor influencing relative consent. Similarly, age (P = .02, >50 years: OR = 7.55, 95% CI: 1.21-47.25) was the only factor influencing willingness to donate before death.In conclusion, specific demographic factors could indicate DPCDs and might reveal directions and methods for cornea donation coordination in the future.

ELECTRICAL SIGNALING IN CONTROL OF OCULAR CELL BEHAVIORS

PubMed Central

Zhao, Min; Chalmers, Laura; Cao, Lin; Viera, Ana C.; Mannis, Mark; Reid, Brian

2011-01-01

Epithelia of the cornea, lens and retina contain a vast array of ion channels and pumps. Together they produce a polarized flow of ions in and out of cells, as well as across the epithelia. These naturally occurring ion fluxes are essential to the hydration and metabolism of the ocular tissues, especially for the avascular cornea and lens. The directional transport of ions generates electric fields and currents in those tissues. Applied electric fields affect migration, division and proliferation of ocular cells which are important in homeostasis and healing of the ocular tissues. Abnormalities in any of those aspects may underlie many ocular diseases, for example chronic corneal ulcers, posterior capsule opacity after cataract surgery, and retinopathies. Electric field-inducing cellular responses, termed electrical signaling here, therefore may be an unexpected yet powerful mechanism in regulating ocular cell behavior. Both endogenous electric fields and applied electric fields could be exploited to regulate ocular cells. We aim to briefly describe the physiology of the naturally occurring electrical activities in the corneal, lens, and retinal epithelia, to provide experimental evidence of the effects of electric fields on ocular cell behaviors, and to suggest possible clinical implications. PMID:22020127

BALB/c and C57BL6 mouse strains vary in their ability to heal corneal epithelial debridement wounds

PubMed Central

Pal-Ghosh, Sonali; Tadvalkar, Gauri; Jurjus, Rosalyn A.; Zieske, James D.; Stepp, Mary Ann

2008-01-01

Genetically engineered mice are usually produced on a mixed genetic background and can be derived from several mouse strains including 129SvJ, C57BL6, and BALB/c. To determine whether differences in recurrent corneal epithelial erosions (RCEEs), corneal epithelial stem cell deficiency (CESCD), and cell migration rate vary between two different mouse strains (BALB/c and C57BL6), 8 week mice were subjected to 1.5 (small) or 2.8 mm (large) manual debridement wounds and allowed to heal for 4 weeks. Syndecan-1 (sdc-1) null mice backcrossed seven generations onto a BALB/c genetic background were also included in the RCEE and CESCD studies to permit comparisons between genotypes within a single strain. After sacrifice, corneas were assessed for the presence of recurrent erosions; no fewer than 15 corneas were used for each strain or genotype studied. Data show that the frequency of recurrent erosions after small wounds was 81 +/− 9% in the C57BL6 mice, 73 +/− 2% in the BALB/c mice, and 32 +/− 6% in sdc-1 null mice. Neither strain developed CESCD after small wounds. The frequency of erosions after large wounds was greater (88 +/− 8%) in the C57BL6 mice compared to BALB/c (60 +/− 2%), and sdc-1 null mice (32 +/− 5%). 4 weeks after the large wounds, fixed, flat mounted corneas were assessed for evidence of CESCD with antibodies against the conjunctival keratin K8 and the goblet cell marker, the mucin Muc5AC. The frequency of CESCD 4 weeks after the large wounds was significantly greater in the C57BL6 mice than in the BALB/c or sdc-1 null mice. To assess cell migration rates, corneas were subjected to 1.5 mm wounds and allowed to heal for 12, 15, 18, 21, and 24 hours. After sacrifice, corneas were stained with Richardson stain (BALB/c) or propidium iodide (C57BL6) to assess reepithelialization rates. While reepithelialization rates were similar for the early times after wounding, by 24 hours the C57BL6 corneas had healed faster: 16 of 30 corneas from the C57BL6 mice were closed compared to 9 of 30 of the BALB/c wounds. BALB/c corneas appeared larger overall compared to C57BL6 corneas; measurements of the overall mass of the enucleated eyes and diameters of the flat-mounted corneas confirmed that C57BL6 eyes and corneas were 6.8% and 4.4% smaller respectively than those of BALB/c mice even though the masses of the two mouse strains at 8 weeks of age were identical. Using BrdU to label dividing cells, we found that 18 hours after wounding, C57BL6 and BALB/c corneal epithelia showed similar numbers of proliferating cells. To determine if the enhanced corneal epithelial cell migration rate seen in the C57BL6 mice was specific to the cornea, we conducted time-lapse studies to assess random cell migration rates in vitro using primary cultures of mouse epidermal keratinocytes. Consistent with the in vivo data, epidermal keratinocytes derived from BALB/c mice migrated 60% slower than C57BL6 cells. These data prove that strain-specific differences in cell migration rate in vivo are present in the cornea and are accompanied by differences in the frequencies of recurrent erosions and corneal epithelial stem cell deficiency. PMID:18809399

Standardized DMEK Technique: Reducing Complications Using Prestripped Tissue, Novel Glass Injector, and Sulfur Hexafluoride (SF6) Gas.

PubMed

Terry, Mark A; Straiko, Michael D; Veldman, Peter B; Talajic, Julia C; VanZyl, Cor; Sales, Christopher S; Mayko, Zachary M

2015-08-01

To report low complication rates in Descemet membrane endothelial keratoplasty (DMEK) using sulfur hexafluoride (SF6) gas, a novel glass injector, and donor tissue prestripped by an eye bank technician. A standardized technique of DMEK was performed in 80 consecutive Fuchs corneal dystrophy cases using technician-prestripped tissue, a novel glass injector, a modified Yoeruek tap technique, and an SF6 gas (20% concentration) bubble for prolonged tissue support. Twenty-five donors were premarked with an "S" stamp for intraoperative orientation. Surgery was performed by 2 experienced DMEK surgeons and 2 inexperienced cornea fellows. Complications were recorded, and the percent endothelial cell loss was calculated at 6 months postoperatively. There were 5 cases that received an air bubble injection postoperatively (6% rebubble rate). There were 6 grafts that immediately failed, 2 because of excessive surgical trauma, and 4 because of upside-down graft placement documented by optical coherence tomography. None of the 25 cases with an S stamp failed. Recipient corneas cleared quickly with no clinical evidence of toxicity from the SF6 gas bubble, and the grafts experienced a mean endothelial cell loss of 27% at 6 months. Tissue prestripped by an eye bank technician can be safely used for DMEK surgery. SF6 gas for prolonged tissue support may reduce the rebubble rate in DMEK, with no apparent acute toxic effect. An unrecognized upside-down graft was the primary cause of graft failure in this series. Upside-down grafts may be eliminated by the use of donor tissue premarked by the eye bank with an S orientation stamp.

Development and Implementation of a Transversely Isotropic Hyperelastic Constitutive Model With Two Fiber Families to Represent Anisotropic Soft Biological Tissues

DTIC Science & Technology

2014-06-01

brain tissue and skeletal muscles , is also discussed. transversely isotropic hyperelastic, two fiber families, nearly incompressible, anisotropic...comprised of fibrous structures, such as muscles , ligaments, tendons, intervertebral discs and the brain, often exhibit strong anisotropy along these fiber ...directions, e.g., collagen fibers of the cornea, striated muscle fibers in skeletal muscles , multiple axonal directions within the brain. In each case

In situ ultrahigh-resolution optical coherence tomography characterization of eye bank corneal tissue processed for lamellar keratoplasty.

PubMed

Brown, Jamin S; Wang, Danling; Li, Xiaoli; Baluyot, Florence; Iliakis, Bernie; Lindquist, Thomas D; Shirakawa, Rika; Shen, Tueng T; Li, Xingde

2008-08-01

To use optical coherence tomography (OCT) as a noninvasive tool to perform in situ characterization of eye bank corneal tissue processed for lamellar keratoplasty. A custom-built ultrahigh-resolution OCT (UHR-OCT) was used to characterize donor corneal tissue that had been processed for lamellar keratoplasty. Twenty-seven donor corneas were analyzed. Four donor corneas were used as controls, whereas the rest were processed into donor corneal buttons for lamellar transplantation by using hand dissection, a microkeratome, or a femtosecond laser. UHR-OCT was also used to noninvasively characterize and monitor the viable corneal tissue immersed in storage medium over 3 weeks. The UHR-OCT captured high-resolution images of the donor corneal tissue in situ. This noninvasive technique showed the changes in donor corneal tissue morphology with time while in storage medium. The characteristics of the lamellar corneal tissue with each processing modality were clearly visible by UHR-OCT. The in situ characterization of the femtosecond laser-cut corneal tissue was noted to have more interface debris than shown by routine histology. The effects of the femtosecond laser microcavitation bubbles on the corneal tissue were well visualized at the edges of the lamellar flap while in storage medium. The results of our feasibility study show that UHR-OCT can provide superb, in situ microstructural characterization of eye bank corneal tissue noninvasively. The UHR-OCT interface findings and corneal endothelial disc thickness uniformity analysis are valuable information that may be used to optimize the modalities and parameters for lamellar tissue processing. The UHR-OCT is a powerful approach that will allow us to further evaluate the tissue response to different processing techniques for posterior lamellar keratoplasty. It may also provide information that can be used to correlate with postoperative clinical outcomes. UHR-OCT has the potential to become a routine part of tissue analysis for any eye bank or centers creating customized lamellar corneal tissue for transplantation.

21 CFR 886.4370 - Keratome.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Keratome. 886.4370 Section 886.4370 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES... battery-powered device intended to shave tissue from sections of the cornea for a lamellar (partial...

Corneal Tissue From Dry Eye Donors Leads to Enhanced Graft Rejection.

PubMed

Inomata, Takenori; Hua, Jing; Nakao, Takeshi; Shiang, Tina; Chiang, Homer; Amouzegar, Afsaneh; Dana, Reza

2018-01-01

To assess the effect of dry eye disease (DED) in graft donors on dendritic cell (DC) maturation, host T-cell sensitization, and corneal allograft rejection. Corneas of control (healthy donor) and DED mice (C57BL/6) were transplanted onto fully allogeneic naive BALB/c recipients (n = 10 mice/group). Long-term allograft survival was evaluated for 8 weeks. Corneas and draining lymph nodes (dLNs) were harvested at posttransplantation day 14 (n = 5 mice/group). The frequencies of MHCII CD11c DCs in the donor corneas and host dLNs and the frequencies of interferon (IFN)-γ and IL-17 CD4 T cells and Foxp3 expression by Tregs in host dLNs were investigated using flow cytometry. The enzyme-linked immunospot assay was used to assess host T-cell allosensitization through direct and indirect pathways (n = 3/group). Recipients of DED donor corneas showed significantly reduced graft survival (10%) compared with control mice (50% survival, P = 0.022), and had significantly increased frequencies of mature DCs in the grafted cornea (DED donor 44.0% ± 0.36% vs. healthy donor 35.4 ± 0.5%; P < 0.0001) and host dLNs (DED donor 25.1% ± 0.66% vs. healthy donor 13.7% ± 1.6%; P = 0.005). Frequencies of IFN-γ and IL-17 T cells were increased in the dLNs of recipients of DED corneas, whereas the expression (mean fluorescence intensity) of Foxp3 in Tregs was decreased significantly in these mice (DED donor 6004 ± 193 vs. healthy donor 6806 ± 81; P = 0.0002). Enzyme-linked immunospot analysis showed that the direct pathway of allosensitization was significantly amplified in recipients of grafts with DED (P = 0.0146). Our results indicate that DED in the donor is a significant risk factor for subsequent corneal allograft rejection.

Cornea Preservation Time Study: Methods and Potential Impact on the Cornea Donor Pool in the United States

PubMed Central

Szczotka-Flynn, Loretta B.; Ayala, Allison R.; Benetz, Beth A.; Gal, Robin L.; Aldave, Anthony J.; Corrigan, Michelle M.; Dunn, Steven P.; McCall, Ty L.; Pramanik, Sudeep; Rosenwasser, George O.; Ross, Kevin W.; Terry, Mark A.; Verdier, David D.

2015-01-01

Purpose: The aim of this study was to describe the aims, methods, donor and recipient cohort characteristics, and potential impact of the Cornea Preservation Time Study (CPTS). Methods: The CPTS is a randomized clinical trial conducted at 40 clinical sites (70 surgeons) designed to assess the effect of donor cornea preservation time (PT) on graft survival 3 years after Descemet stripping automated endothelial keratoplasty (DSAEK). Eyes undergoing surgery for Fuchs endothelial corneal dystrophy or pseudophakic/aphakic corneal edema were randomized to receive donor corneas stored ≤7 days or 8 to 14 days. Donor and patient characteristics, tissue preparation and surgical parameters, recipient and donor corneal stroma clarity, central corneal thickness, intraocular pressure, complications, and a reading center-determined central endothelial cell density were collected. Surveys were conducted to evaluate pre-CPTS PT practices. Results: The 1330 CPTS donors were: 49% >60 years old, 27% diabetic, had a median eye bank–determined screening endothelial cell density of 2688 cells/mm2, and 74% eye bank prepared for DSAEK. A total of 1090 recipients (1330 eyes including 240 bilateral cases) had: median age of 70 years, were 60% female, 90% white, 18% diabetic, 52% phakic, and 94% had Fuchs endothelial corneal dystrophy. Before the CPTS, 19 eye banks provided PT data on 20,852 corneas domestically placed for DSAEK in 2010 to 2011; 96% were preserved ≤7 days. Of 305 American Academy of Ophthalmology members responding to a pre-CPTS survey, 233 (76%) set their maximum PT preference at 8 days or less. Conclusions: The CPTS will increase understanding of factors related to DSAEK success and, if noninferiority of longer PT is shown, will have great potential to extend the available pool of endothelial keratoplasty donors. Clinical Trial Registration—URL: http://www.clinicaltrials.gov. Unique identifier: NCT01537393. PMID:25850706

Synthetic versus serum-based medium for corneal preservation in organ culture: a comparative study between 2 different media.

PubMed

Parekh, Mohit; Ferrari, Stefano; Salvalaio, Gianni; Ponzin, Diego

2015-01-01

To study the effect of a synthetic medium and compare it with a serum-based medium for corneal preservation in organ culture using an overall quality assessment system. A randomized study with blinded observers was performed comparing parameters such as thickness, transparency, viable endothelial cell density (VECD), morphology, and overall quality (OQ) of the corneal tissues preserved in synthetic and a serum-based medium, respectively. Seven human paired corneas were randomly selected and assessed at day 0 (initial), day 2 (before organ culture), day 30 (before deturgescence/deswelling storage), and 48 hours post deswelling. Thickness was determined with optical coherence tomography and transparency with a validated, custom device. The morphology and VECD were observed after treating the tissues with trypan blue and sucrose. Data were compared using paired t tests with p<0.05 deemed significant. Parameters were similar at the initial stage between the groups with no statistically significant difference. However, after preservation in the deturgescent medium, the corneas stored in a serum-based medium showed a higher and statistically significant OQ value (p = 0.0317). The OQ of a serum-based medium was higher than that of the synthetic medium. A higher rate of transparency and reduction in thickness was observed in the serum-based medium at the end of the storage. Although complete synthetic media may have distinct advantages of being serum/animal-free, the quality of the cornea is of a reasonable concern when it is deemed for transplantation.

Transcription, Translation, and Function of Lubricin, a Boundary Lubricant, at the Ocular Surface

PubMed Central

Schmidt, Tannin A.; Sullivan, David A.; Knop, Erich; Richards, Stephen M.; Knop, Nadja; Liu, Shaohui; Sahin, Afsun; Darabad, Raheleh Rahimi; Morrison, Sheila; Kam, Wendy R.; Sullivan, Benjamin D.

2013-01-01

Importance Lubricin may be an important barrier to the development of corneal and conjunctival epitheliopathies that may occur in dry eye disease and contact lens wear. Objective To test the hypotheses that lubricin (ie, proteoglycan 4 [PRG4]), a boundary lubricant, is produced by ocular surface epithelia and acts to protect the cornea and conjunctiva against significant shear forces generated during an eyelid blink and that lubricin deficiency increases shear stress on the ocular surface and promotes corneal damage. Design, Setting, and Participants Human, porcine, and mouse tissues and cells were processed for molecular biological, immunohistochemical, and tribological studies, and wild-type and PRG4 knockout mice were evaluated for corneal damage. Results Our findings demonstrate that lubricin is transcribed and translated by corneal and conjunctival epithelial cells. Lubricin messenger RNA is also present in lacrimal and meibomian glands, as well as in a number of other tissues. Absence of lubricin in PRG4 knockout mice is associated with a significant increase in corneal fluorescein staining. Our studies also show that lubricin functions as an effective friction-lowering boundary lubricant at the human cornea-eyelid interface. This effect is specific and cannot be duplicated by the use of hyaluronate or bovine serum albumin solutions. Conclusions and Relevance Our results show that lubricin is transcribed, translated, and expressed by ocular surface epithelia. Moreover, our findings demonstrate that lubricin presence significantly reduces friction between the cornea and conjunctiva and that lubricin deficiency may play a role in promoting corneal damage. PMID:23599181

Transcription, translation, and function of lubricin, a boundary lubricant, at the ocular surface.

PubMed

Schmidt, Tannin A; Sullivan, David A; Knop, Erich; Richards, Stephen M; Knop, Nadja; Liu, Shaohui; Sahin, Afsun; Darabad, Raheleh Rahimi; Morrison, Sheila; Kam, Wendy R; Sullivan, Benjamin D

2013-06-01

Lubricin may be an important barrier to the development of corneal and conjunctival epitheliopathies that may occur in dry eye disease and contact lens wear. To test the hypotheses that lubricin (ie, proteoglycan 4 [PRG4 ]), a boundary lubricant, is produced by ocular surface epithelia and acts to protect the cornea and conjunctiva against significant shear forces generated during an eyelid blink and that lubricin deficiency increases shear stress on the ocular surface and promotes corneal damage. Human, porcine, and mouse tissues and cells were processed for molecular biological, immunohistochemical, and tribological studies, and wild-type and PRG4 knockout mice were evaluated for corneal damage. Our findings demonstrate that lubricin is transcribed and translated by corneal and conjunctival epithelial cells. Lubricin messenger RNA is also present in lacrimal and meibomian glands, as well as in a number of other tissues. Absence of lubricin in PRG4 knockout mice is associated with a significant increase in corneal fluorescein staining. Our studies also show that lubricin functions as an effective friction-lowering boundary lubricant at the human cornea-eyelid interface. This effect is specific and cannot be duplicated by the use of hyaluronate or bovine serum albumin solutions. Our results show that lubricin is transcribed, translated, and expressed by ocular surface epithelia. Moreover, our findings demonstrate that lubricin presence significantly reduces friction between the cornea and conjunctiva and that lubricin deficiency may play a role in promoting corneal damage.

Disruption of epithelial cell migration as a potential mechanism of cleft palate induction

EPA Science Inventory

Cleft palate occurs in about one in seven hundred births per year, making it the most prevalent craniofacial birth defect in the world. During embryonic development, tissue fusion is a critical step in the formation of the palate, cornea, urethra, and neural tube. Epithelial cell...

Biocompatibility and light transmission of liposomal lenses.

PubMed

Danion, Anne; Doillon, Charles J; Giasson, Claude J; Djouahra, Saliha; Sauvageau, Patrick; Paradis, Renée; Vermette, Patrick

2007-10-01

To validate the biocompatibility and transmittance properties of contact lenses bearing intact liposomes. These liposomal lenses loaded with therapeutics can be used as ophthalmic drug delivery systems. The biocompatibility of soft contact lenses, coated with liposomes was evaluated through in vitro direct and indirect cytocompatibility assays on human corneal epithelial cells, on reconstructed human corneas and on ex vivo rabbit corneas. The direct and indirect transmission spectra of liposome-covered lenses were also evaluated to test if they transmit all wavelengths of the ultraviolet-visible spectrum, to thereby fulfill their optical function, without gross alteration of the colors perception and with a minimum of light dispersion. Contact lenses bearing layers of stable liposomes did not induce any significant changes in cell viability and in cell growth, compared with lenses bearing no liposome. Elution assays revealed that no cytotoxic compound leaks from the lenses whether bearing liposomes or not. Histological analyses of reconstructed human corneas and ex vivo rabbit corneas directly exposed to liposomal lenses revealed neither alteration to the cell nor to the tissue structures. Contact lenses bearing layers of liposomes did not significantly affect light transmission compared with control lenses without liposome at the wavelength of maximal photopic sensitivity, i.e., 550 nm. In addition, the contact lenses afford more eye protection in the ultraviolet spectrum, compared with the control lenses. Liposomal contact lenses are biocompatible and their transmittance properties are not affected in the visible light range.

Quantifying the effects of hydration on corneal stiffness with optical coherence elastography

NASA Astrophysics Data System (ADS)

Singh, Manmohan; Li, Jiasong; Han, Zhaolong; Vantipalli, Srilatha; Aglyamov, Salavat R.; Twa, Michael D.; Larin, Kirill V.

2018-02-01

Several methods have been proposed to assess changes in corneal biomechanical properties due to various factors, such as degenerative diseases, intraocular pressure, and therapeutic interventions (e.g. corneal collagen crosslinking). However, the effect of the corneal tissue hydration state on corneal stiffness is not well understood. In this work, we induce low amplitude (< 10 μm) elastic waves with a focused micro air-pulse in fresh in situ rabbit corneas (n = 10) in the whole eye-globe configuration at an artificially controlled intraocular pressure. The waves were then detected with a phase-stabilized swept source optical coherence elastography system. Baseline measurements were taken every 20 minutes for an hour while the corneas were hydrated with 1X PBS. After the measurement at 60 minutes, a 20% dextran solution was topically instilled to dehydrate the corneas. The measurements were repeated every 20 minutes again for an hour. The results showed that the elastic wave velocity decreased as the corneal thickness decreased. Finite element modeling (FEM) was performed using the corneal geometry and elastic wave propagation speed to assess the stiffness of the samples. The results show that the stiffness increased from 430 kPa during hydration with PBS to 500 kPa after dehydration with dextran, demonstrating that corneal hydration state, apart from geometry and intraocular pressure, can change the stiffness of the cornea.

Pharmacokinetics of pirfenidone after topical administration in rabbit eye

PubMed Central

Sun, Guoying; Lin, Xianchai; Zhong, Hua; Yang, Yangfan; Qiu, Xuan; Ye, Chengtian; Wu, Kaili

2011-01-01

Purpose Pirfenidone (5-methyl-1-phenyl-2-[1H]-pyridone) is a new, broad-spectrum agent that has an inhibition effect on the proliferation, migration, and collagen contraction of human Tenon’s fibroblasts, and thus modulating the wound healing process of glaucoma filtering surgical site. This study investigated the pharmacokinetics of topically administered pirfenidone (0.5%) in rabbit eyes. Methods Pirfenidone solution (50 μl) was instilled into the rabbit’s conjunctival sac. The rabbits were quickly sacrificed at 2, 5, 8, 10, 15, 20, 30, 60, 90, and 120 min after the administration and ocular tissues were obtained. The concentrations of pirfenidone in conjunctiva, sclera, cornea, aqueous humor, and vitreous were determined by high performance liquid chromatography. Results After topical administration, there was wide distribution and fast clearance of pirfenidone among the various ocular tissues. The mean maximum concentrations (Cmax) of pirfenidone in cornea, conjunctiva, sclera, aqueous humor, and vitreous were 9.64 mg/g, 9.62 mg/g, 2.13 mg/g, 34.88 mg/l and 0.52 mg/l, respectively. The half-life for these tissues was 18.26, 34.16, 15.71, 70.91, and 39.48 min, respectively. Conclusions Measurable concentrations of pirfenidone are achieved in ocular tissues after topical application in rabbit model. Topical administration of pirfenidone may be an effective approach for modulation of wound healing responses in glaucoma filtration surgical site. PMID:21866212

Indentation and needle insertion properties of the human eye

PubMed Central

Matthews, A; Hutnik, C; Hill, K; Newson, T; Chan, T; Campbell, G

2014-01-01

Purpose Characterization of the biomechanical properties of the human eye has a number of potential utilities. One novel purpose is to provide the basis for development of suitable tissue-mimicking material. The purpose of this study was to determine the indentation and needle insertion characteristics on human eye globes and tissue strips. Methods An indenter assessed the elastic response of human eye globes and tissue strips under increasing compressive loads. Needle insertion determined the force (N) needed to penetrate various areas of the eye wall. Results The results demonstrated that globes underwent slightly greater indentation at the midline than at the central cornea, and corneal strips indented twofold more than scleral strips, although neither difference was significant (P=0.400 and P=0.100, respectively). Significant differences were observed among various areas of needle insertion (P<0.001). Needle insertion through the anterior sclera (adjacent to the limbus) and posterior sclera (adjacent to the optic nerve) required the greatest amount of force (0.954 and 1.005 N, respectively). The force required to penetrate the central cornea (0.518 N) was significantly lower than all other areas except the midline sclera (0.700 N) Conclusion These data form the basis for further research into the development of a tissue-mimicking human eye construct with potential utility as a model for use in ophthalmology research and surgical teaching. PMID:24810571

Research on the comparison of the demethylvancomycin's diffusion-deposition characteristics in the ocular solid tissues of sustained subtenon drug delivery with subconjunctival injection.

PubMed

Duan, Yi-Qin; Yang, Ye-Zhen; Huang, Xue-Tao; Lin, Ding

2017-11-01

To compare the demethylvancomycin's diffusion-deposition characteristics in the ocular solid tissues of sustained subtenon drug delivery with subconjunctival injection. Sixty adult white rabbits were randomly assigned to the subtenon drug delivery group and the subconjunctival injection group. The subtenon drug delivery group was continuously infused demethylvancomycin to the subtenon of rabbits. The subconjunctival injection group was injected demethylvancomycin to the subconjunctival of rabbits. Cornea, iris and sclera were collected for high-performance liquid chromatography analyses to determine drug concentrations at one hour, three hours, six hours, 12 h and 24 h of drug administration. WinNonlin 6.3 was used to calculate the parameters of cumulative area under the curve (AUC cum ) of demethylvancomycin. The peak levels of demethylvancomycin concentration of the subtenon drug delivery group and the subconjunctival injection group were 92.406 ± 21.555 and 51.778 ± 14.001 μg/g in cornea, 28.451 ± 10.229 μg/g and 42.271 ± 27.291 μg/g in iris, 153.166 ± 51.738 μg/g and 57.423 ± 18.480 μg/g in sclera. The differences of concentrations between the two groups in cornea and sclera were statistically significant (F = 487.775, p < 0.001; F = 132.748, p < 0.001). The difference in iris was not statistically significant (F = 4.848, p = 0.064). The maximum of AUC cum of the subtenon drug delivery group and the subconjunctival injection group was 1808.23 h * μg/g and 273.73 h * μg/g in cornea, 489.12 h * μg/g and 216.16 h * μg/g in iris and 2166.34 h * μg/g and 392.57 h * μg/g in sclera at 24 h of drug administration. The sustained subtenon drug delivery had a better drug permeability and accumulation in the intraocular solid tissue compared to subconjunctival injection, which demonstrated it was probably a promising and effective approach for treating posterior segment diseases and endophthalmitis.

Molecular crowding of collagen: a pathway to produce highly-organized collagenous structures.

PubMed

Saeidi, Nima; Karmelek, Kathryn P; Paten, Jeffrey A; Zareian, Ramin; DiMasi, Elaine; Ruberti, Jeffrey W

2012-10-01

Collagen in vertebrate animals is often arranged in alternating lamellae or in bundles of aligned fibrils which are designed to withstand in vivo mechanical loads. The formation of these organized structures is thought to result from a complex, large-area integration of individual cell motion and locally-controlled synthesis of fibrillar arrays via cell-surface fibripositors (direct matrix printing). The difficulty of reproducing such a process in vitro has prevented tissue engineers from constructing clinically useful load-bearing connective tissue directly from collagen. However, we and others have taken the view that long-range organizational information is potentially encoded into the structure of the collagen molecule itself, allowing the control of fibril organization to extend far from cell (or bounding) surfaces. We here demonstrate a simple, fast, cell-free method capable of producing highly-organized, anistropic collagen fibrillar lamellae de novo which persist over relatively long-distances (tens to hundreds of microns). Our approach to nanoscale organizational control takes advantage of the intrinsic physiochemical properties of collagen molecules by inducing collagen association through molecular crowding and geometric confinement. To mimic biological tissues which comprise planar, aligned collagen lamellae (e.g. cornea, lamellar bone or annulus fibrosus), type I collagen was confined to a thin, planar geometry, concentrated through molecular crowding and polymerized. The resulting fibrillar lamellae show a striking resemblance to native load-bearing lamellae in that the fibrils are small, generally aligned in the plane of the confining space and change direction en masse throughout the thickness of the construct. The process of organizational control is consistent with embryonic development where the bounded planar cell sheets produced by fibroblasts suggest a similar confinement/concentration strategy. Such a simple approach to nanoscale organizational control of structure not only makes de novo tissue engineering a possibility, but also suggests a clearer pathway to organization for fibroblasts than direct matrix printing. Copyright © 2012 Elsevier Ltd. All rights reserved.

In vivo THz sensing of the cornea of the eye

NASA Astrophysics Data System (ADS)

Ozheredov, Ilya; Prokopchuk, Mikhail; Mischenko, Mikhail; Safonova, Tatiana; Solyankin, Petr; Larichev, Andrey; Angeluts, Andrey; Balakin, Alexei; Shkurinov, Alexander

2018-05-01

Measurement of the absolute value of the humidity of the cornea of the human eye and its dynamics is of paramount importance for the preservation of eyesight. In the present paper we have demonstrated that terahertz technologies can be practically applied for quantitative measurement of the physiological dynamics of tear film and sensing of corneal tissue hydration. We suggest uses of the equipment for application in clinics and a method for absolute calibration of the values for measurement. The proposed method is fundamentally different from existing and currently available methods of ophthalmological diagnosis. This suggests that the developed technique may have high diagnostic significance and can be used in the study and treatment of several diseases of the ocular surface.

Controlled reshaping of the front surface of the cornea through its full-area ablation outside of the optical zone with a Gaussian ArF excimer laser beam

NASA Astrophysics Data System (ADS)

Semchishen, A. V.; Semchishen, V. A.

2014-01-01

We studied in vitro the response of the topography of the cornea to its full-area laser ablation (the laser beam spot diameter is commensurable with the size of the interface) outside of the central zone with an excimer laser having a Gaussian fluence distribution across the beam. Subject to investigation were the topographically controlled surface changes of the anterior cornea in 60 porcine eyes with a 5 ± 1.25-diopter artificially induced astigmatism, the changes being caused by laser ablation of the stromal collagen in two 3.5-mm-dia. circular areas along the weaker astigmatism axis. Experimental relationships are presented between the actual astigmatism correction and the expected correction for the intact optical zones 1, 2, 3, and 4 mm in diameter. The data for each zone were approximated by the least-squares method with the function d = a + bx. The coefficient b is given with the root-mean-square error. The statistical processing of the data yielded the following results: d = (0.14 ± 0.037)x for the 1-mm-dia. optical zone, (1.10 ± 0.036)x for the 2-mm-dia. optical zone, (1.04 ± 0.020)x for the 3-mm-dia. optical zone, and (0.55 ± 0.04)x for the 4-mm-dia. optical zone. Full astigmatism correction was achieved with ablation effected outside of the 3-mm-dia. optical zone. The surface changes of the cornea are shown to be due not only to the removal of the corneal tissue, but also to the biomechanical topographic response of the cornea to its strain caused by the formation of a dense pseudomembrane in the ablation area.

Effects of topical nonsteroidal antiinflammatory drugs on the expression of matrix metalloproteinases in the cornea.

PubMed

Reviglio, Victor E; Rana, Tayyib S; Li, Qian J; Ashraf, M Farooq; Daly, Mary K; O'Brien, Terrence P

2003-05-01

To assess the effects of nonsteroidal antiinflammatory drug (NSAID) eyedrops on the expression of matrix metalloproteinases in corneal tissue. Ocular Microbiology and Immunology Laboratory, Refractive Surgery Research Laboratory, The Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, USA. Seventy rats were divided into 2 groups: intact and debrided epithelium. Uniform central corneal epithelial defects were created in the right eye of the debrided corneal group. Each group was divided into 4 subgroups, each receiving 1 of the following eyedrops or artificial tears: The 3 NSAIDs were diclofenac sodium 0.1% (Falcon or Voltaren) and preservative-free ketorolac 0.5% (Acular PF). The artificial tears were carboxymethylcellulose sodium 0.5% (Refresh Plus PF). The eyedrops were administered 4 times a day for 1 week. The rats were killed on days 2 and 7. The corneas were excised and processed for immunohistochemical staining, Western blot assay, and zymography studies to determine the localization of the production of the following matrix metalloproteinases (MMPs): MMP-1, MMP-2, MMP-8, and MMP-9. Matrix metalloproteinase-1, MMP-8, and MMP-2 were detected in rat corneas at 48 hours in the debrided and intact epithelium groups treated with NSAID eyedrops. The MMP-1 and MMP-8 expression levels were higher in intact corneas in the diclofenac sodium groups than in the ketorolac and artificial tears groups. The expression was localized mostly in the epithelial cells and occasionally in keratocytes. This study provides preliminary evidence that topical application of some NSAIDs can induce the early expression of MMP-1, MMP-2, and MMP-8 in the cornea, suggesting that MMPs play a role in the corneal cytotoxicity of certain NSAIDs.

HIV and hepatitis C virus RNA in seronegative organ and tissue donors.

PubMed

Challine, Dominique; Pellegrin, Bertrand; Bouvier-Alias, Magali; Rigot, Pierrette; Laperche, Liliane; Pawlotsky, Jean-Michel

The objective of our study was to determine whether nucleic acid testing could detect HIV RNA or hepatitis C virus (HCV) RNA in a large series of seronegative organ and tissue donors, and whether this technique should be routinely used to improve viral safety of grafts. We studied 2236 organ donors, 636 tissue donors, and 177 cornea donors. We identified five HCV RNA-positive donors in 2119 HCV-seronegative organ donors, and one HCV RNA-positive donor in 631 HCV-seronegative tissue donors. No HIV-seronegative, HIV RNA-positive donor was identified. Our data suggest that routine nucleic acid testing of organ and tissue donors might increase viral safety in transplantation.

Moxifloxacin: Clinically compatible contrast agent for multiphoton imaging

NASA Astrophysics Data System (ADS)

Wang, Taejun; Jang, Won Hyuk; Lee, Seunghun; Yoon, Calvin J.; Lee, Jun Ho; Kim, Bumju; Hwang, Sekyu; Hong, Chun-Pyo; Yoon, Yeoreum; Lee, Gilgu; Le, Viet-Hoan; Bok, Seoyeon; Ahn, G.-One; Lee, Jaewook; Gho, Yong Song; Chung, Euiheon; Kim, Sungjee; Jang, Myoung Ho; Myung, Seung-Jae; Kim, Myoung Joon; So, Peter T. C.; Kim, Ki Hean

2016-06-01

Multiphoton microscopy (MPM) is a nonlinear fluorescence microscopic technique widely used for cellular imaging of thick tissues and live animals in biological studies. However, MPM application to human tissues is limited by weak endogenous fluorescence in tissue and cytotoxicity of exogenous probes. Herein, we describe the applications of moxifloxacin, an FDA-approved antibiotic, as a cell-labeling agent for MPM. Moxifloxacin has bright intrinsic multiphoton fluorescence, good tissue penetration and high intracellular concentration. MPM with moxifloxacin was demonstrated in various cell lines, and animal tissues of cornea, skin, small intestine and bladder. Clinical application is promising since imaging based on moxifloxacin labeling could be 10 times faster than imaging based on endogenous fluorescence.

Transporter targeted gatifloxacin prodrugs: Synthesis, permeability, and topical ocular delivery

PubMed Central

Vooturi, Sunil K.; Kadam, Rajendra S.; Kompella, Uday B.

2013-01-01

Purpose To design and synthesize prodrugs of gatifloxacin targeting OCT, MCT, and ATB (0, +) transporters and to identify a prodrug with enhanced delivery to the back of the eye. Method Dimethylamino-propyl, carboxy-propyl, and amino-propyl(2-methyl) derivatives of gatifloxacin (GFX), DMAP-GFX, CP-GFX, and APM-GFX, were designed and synthesized to target OCT, MCT, and ATB (0, +) transporters, respectively. LC-MS method was developed to analyze drug and prodrug levels in various studies. Solubility and Log D (pH 7.4) were measured for prodrugs and the parent drug. Permeability of the prodrugs was determined in cornea, conjunctiva, and sclera-choroidretinal pigment epitheluim (SCRPE) and compared with gatifloxacin using Ussing chamber assembly. Permeability mechanisms were elucidated by determining the transport in the presence of transporter specific inhibitors. 1-Methyl-4-phenylpyridinium iodide (MPP+), nicotinic acid sodium salt, and α-methyl-DL-tryptophan were used to inhibit OCT, MCT, and ATB (0, +) transporters, respectively. A prodrug selected based on in vitro studies was administered as an eye drop to pigmented rabbits and the delivery to various eye tissues including vitreous humor was compared with gatifloxacin dosing. Results DMAP-GFX exhibited 12.8-fold greater solubility than GFX. All prodrugs were more lipophilic, with the measured Log D (pH 7.4) values ranging from 0.05 to 1.04, when compared to GFX (Log D: -1.15). DMAP-GFX showed 1.4-, 1.8-, and 1.9-fold improvement in permeability across cornea, conjunctiva, as well as SCRPE when compared to GFX. Moreover, it exhibited reduced permeability in the presence of MPP+ (competitive inhibitor of OCT), indicating OCT-mediated transport. CP-GFX showed 1.2-, 2.3- and 2.5-fold improvement in permeability across cornea, conjunctiva and SCRPE, respectively. In the presence of nicotinic acid (competitive inhibitor of MCT), permeability of CP-GFX was reduced across conjunctiva. However, cornea and SCRPE permeability of CP-GFX was not affected by nicotinic acid. APM-GFX did not show any improvement in permeability when compared to GFX across cornea, conjunctiva, and SCRPE. Based on solubility and permeability, DMAP-GFX was selected for in vivo studies. DMAP-GFX showed 3.6- and 1.95-fold higher levels in vitreous humor and CRPE compared to that of GFX at 1 hour after topical dosing. In vivo conversion of DMAP-GFX prodrug to GFX was quantified in tissues isolated at 1 hour after dosing. Prodrug-to-parent drug ratio was 8, 70, 24, 21, 29, 13, 55, and 60 % in cornea, conjunctiva, iris-ciliary body, aqueous humor, sclera, CRPE, retina, and vitreous humor, respectively. Conclusions DMAP-GFX prodrug enhanced solubility, Log D, as well as OCT mediated delivery of gatifloxacin to the back of the eye. PMID:23003105

Finesse of transparent tissue cutting by ultrafast lasers at various wavelengths.

PubMed

Wang, Jenny; Schuele, Georg; Palanker, Daniel

2015-01-01

Transparent ocular tissues, such as the cornea and crystalline lens, can be ablated or dissected using short-pulse lasers. In refractive and cataract surgeries, the cornea, lens, and lens capsule can be cut by producing dielectric breakdown in the focus of a near-infrared (IR) femtosecond laser, which results in explosive vaporization of the interstitial water, causing mechanical rupture of the surrounding tissue. Here, we compare the texture of edges of lens capsule cut by femtosecond lasers with IR and ultraviolet (UV) wavelengths and explore differences in interactions of these lasers with biological molecules. Scanning electron microscopy indicates that a 400-nm laser is capable of producing very smooth cut edges compared to 800 or 1030 nm at a similar focusing angle. Using gel electrophoresis and liquid chromatography/mass spectrometry, we observe laser-induced nonlinear breakdown of proteins and polypeptides by 400-nm femtosecond pulses above and below the dielectric breakdown threshold. On the other hand, 800-nm femtosecond lasers do not produce significant dissociation even above the threshold of dielectric breakdown. However, despite this additional interaction of UV femtosecond laser with proteins, we determine that efficient cutting requires plasma-mediated bubble formation and that remarkably smooth edges are the result of reduced thresholds and smaller focal volume.

THz and mm-Wave Sensing of Corneal Tissue Water Content: Electromagnetic Modeling and Analysis

PubMed Central

Taylor, Zachary D.; Garritano, James; Sung, Shijun; Bajwa, Neha; Bennett, David B.; Nowroozi, Bryan; Tewari, Priyamvada; Sayre, James; Hubschman, Jean-Pierre; Deng, Sophie; Brown, Elliott R.; Grundfest, Warren S.

2015-01-01

Terahertz (THz) spectral properties of human cornea are explored as a function of central corneal thickness (CCT) and corneal water content, and the clinical utility of THz-based corneal water content sensing is discussed. Three candidate corneal tissue water content (CTWC) perturbations, based on corneal physiology, are investigated that affect the axial water distribution and total thickness. The THz frequency reflectivity properties of the three CTWC perturbations were simulated and explored with varying system center frequency and bandwidths (Q-factors). The modeling showed that at effective optical path lengths on the order of a wavelength the cornea presents a lossy etalon bordered by air at the anterior and the aqueous humor at the posterior. The simulated standing wave peak-to-valley ratio is pronounced at lower frequencies and its effect on acquired data can be modulated by adjusting the bandwidth of the sensing system. These observations are supported with experimental spectroscopic data. The results suggest that a priori knowledge of corneal thickness can be utilized for accurate assessments of corneal tissue water content. The physiologic variation of corneal thickness with respect to the wavelengths spanned by the THz band is extremely limited compared to all other structures in the body making CTWC sensing unique amongst all proposed applications of THz medical imaging. PMID:26322247

Development of a convenient ex vivo model for the study of the transcorneal permeation of drugs: histological and permeability evaluation.

PubMed

Pescina, Silvia; Govoni, Paolo; Potenza, Arianna; Padula, Cristina; Santi, Patrizia; Nicoli, Sara

2015-01-01

In this paper, an ex vivo model for the study of the transcorneal permeation of drugs, based on porcine tissues, was evaluated. The setup is characterized by ease of realization, absence of O₂ and CO₂ bubbling and low cost; additionally, the large availability of porcine tissue permits a high throughput. Histological images showed the comparability between porcine and human corneas and confirmed the effectiveness of the isolation procedure. A new de-epithelization procedure based on a thermal approach was also set up to simulate cornea permeability in pathological conditions. The procedure did not affect the integrity of the underlying layers and allowed the characterization of the barrier properties of epithelium and stroma. Six compounds with different physicochemical properties were tested: fluorescein, atenolol, propranolol, diclofenac, ganciclovir and lidocaine. The model highlighted the barrier function played by epithelium toward the diffusion of hydrophilic compounds and the permselectivity with regard to more lipophilic molecules. In particular, positively charged compounds showed a significantly higher transcorneal permeability than negatively charged compounds. The comparability of results with literature data supports the goodness and the robustness of the model, especially taking into account the behavior of fluorescein, which is generally considered a marker of tissue integrity. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association.

Lumican as a multivalent effector in wound healing.

PubMed

Karamanou, Konstantina; Perrot, Gwenn; Maquart, Francois-Xavier; Brézillon, Stéphane

2018-03-01

Wound healing, a complex physiological process, is responsible for tissue repair after exposure to destructive stimuli, without resulting in complete functional regeneration. Injuries can be stromal or epithelial, and most cases of wound repair have been studied in the skin and cornea. Lumican, a small leucine-rich proteoglycan, is expressed in the extracellular matrices of several tissues, such as the cornea, cartilage, and skin. This molecule has been shown to regulate collagen fibrillogenesis, keratinocyte phenotypes, and corneal transparency modulation. Lumican is also involved in the extravasation of inflammatory cells and angiogenesis, which are both critical in stromal wound healing. Lumican is the only member of the small leucine-rich proteoglycan family expressed by the epithelia during wound healing. This review summarizes the importance of lumican in wound healing and potential methods of lumican drug delivery to target wound repair are discussed. The involvement of lumican in corneal wound healing is described based on in vitro and in vivo models, with critical emphasis on its underlying mechanisms of action. Similarly, the expression and role of lumican in the healing of other tissues are presented, with emphasis on skin wound healing. Overall, lumican promotes normal wound repair and broadens new therapeutic perspectives for impaired wound healing. Copyright © 2018. Published by Elsevier B.V.

Hazards To The Eye From UV

NASA Astrophysics Data System (ADS)

Zuclich, Joseph A.

1980-10-01

Ocular effects of ultraviolet radiation, 200-400 nm, are reviewed. Depending upon the exposure parameter involved, UV radiation may be harmful to the cornea, lens and/or retina. Ranges of exposure parameters (wavelength, exposure duration, etc.) for which each of the tissues is susceptible are specified and the nature of the tissue is described. Present understanding of the thermal and photochemical damage mechanism operative for various conditions of exposure are discussed Ocular damage thresholds for wide ranges of exposure parameters are summarized and compared to existing safety standards.

Platform for Rapid Delivery of Biologics and Drugs to Ocular Cells and Tissues Following Combat Associated Trauma

DTIC Science & Technology

2013-09-01

death pathways such as apoptosis subsequent to acute trauma as soon as possible, ideally by self- administration of a drug or a biologic that can be... Drugs to Ocular Tissues Including Retina and Cornea . Mol Ther, 2007;16(1):107- 14. 3. Read SP, Cashman SM, and Kumar-Singh R: POD...1 AD_________________ Award Number: W81XWH-12-1-0374 TITLE: Platform for Rapid Delivery of Biologics and Drugs to Ocular Cells

Gamma-Irradiated Sterile Cornea for Use in Corneal Transplants in a Rabbit Model

PubMed Central

Yoshida, Junko; Heflin, Thomas; Zambrano, Andrea; Pan, Qing; Meng, Huan; Wang, Jiangxia; Stark, Walter J.; Daoud, Yassine J.

2015-01-01

Purpose: Gamma irradiated corneas in which the donor keratocytes and endothelial cells are eliminated are effective as corneal lamellar and glaucoma patch grafts. In addition, gamma irradiation causes collagen cross inking, which stiffens collagen fibrils. This study evaluated gamma irradiated corneas for use in corneal transplantations in a rabbit model comparing graft clarity, corneal neovascularization, and edema. Methods: Penetrating keratoplasty was performed on rabbits using four types of corneal grafts: Fresh cornea with endothelium, gamma irradiated cornea, cryopreserved cornea, and fresh cornea without endothelium. Slit lamp examination was performed at postoperative week (POW) one, two, and four. Corneal clarity, edema, and vascularization were graded. Confocal microscopy and histopathological evaluation were performed. A P < 0.05 was statistically significant. Results: For all postoperative examinations, the corneal clarity and edema were statistically significantly better in eyes that received fresh cornea with endothelium compared to the other three groups (P < 0.05). At POW 1, gamma irradiated cornea scored better than the cryopreserved and fresh cornea without endothelium groups in clarity (0.9 vs. 1.5 and 2.6, respectively), and edema (0.6 vs. 0.8 and 2.0, respectively). The gamma irradiated corneas, cryopreserved corneas and the fresh corneas without endothelium, developed haze and edema after POW 2. Gamma irradiated cornea remained statistically significantly clearer than cryopreserved and fresh cornea without endothelium during the observation period (P < 0.05). Histopathology indicated an absence of keratocytes in gamma irradiated cornea. Conclusion: Gamma irradiated corneas remained clearer and thinner than the cryopreserved cornea and fresh cornea without endothelium. However, this outcome is transient. Gamma irradiated corneas are useful for lamellar and patch grafts, but cannot be used for penetrating keratoplasty. PMID:26180475

Coefficient of Friction of Human Corneal Tissue.

PubMed

Wilson, Tawnya; Aeschlimann, Rudolf; Tosatti, Samuele; Toubouti, Youssef; Kakkassery, Joseph; Osborn Lorenz, Katherine

2015-09-01

A novel property evaluation methodology was used to determine the elusive value for the human corneal coefficient of friction (CoF). Using a microtribometer on 28 fresh human donor corneas with intact epithelia, the CoF was determined in 4 test solutions (≥5 corneas/solution): tear-mimicking solution (TMS) in borate-buffered saline (TMS-PS), TMS in phosphate-buffered saline (TMS-PBS), TMS with HEPES-buffered saline (TMS-HEPES), and tear-like fluid in PBS (TLF-PBS). Mean (SD) CoF values ranged from 0.006 to 0.015 and were 0.013 (0.010) in TMS-PS, 0.006 (0.003) in TMS-PBS, 0.014 (0.005) in TMS-HEPES, and 0.015 (0.009) in TLF-PBS. Statistically significant differences were shown for TMS-PBS versus TLF (P = 0.0424) and TMS-PBS versus TMS-HEPES (P = 0.0179), but not for TMS-PBS versus TMS-PS (P = 0.2389). Successful measurement of the fresh human corneal tissue CoF was demonstrated, with values differing in the evaluated buffer solutions, within this limited sample size.

The surface of the eye – a superficial entity with deep repercussions

PubMed Central

Potop, Vasile; Dumitrache, Marieta; Ciocalteu, Alina

2009-01-01

The surface of the eye is an anatomical and functional entity with a relatively recent delimitation but with significant therapeutic and diagnostic consequences. The pathology of the conjunctive and cornea must be approached by looking at the interrelations between the two tissues that are so different anatomically and functionally but in the same time form a unit in structuring the eye’s surface. There are two major categories of relations between the two tissues: one of them is mediated by lachrymal secretion, a process whose complexity is not yet fully understood, and the other is germinal, referring to the stem cells located at the limbus which become epithelial cornea cells that can fixate lachrymal fluid. Imbalances in the quantity and quality of lachrymal secretion can be compensated, up to a certain point, by artificial products, but in severe cases only specially prepared autologous serum can compensate the deficit. The limbic deficits that affect stem cells require complex therapeutic procedures like limbic cell transplant, using an amniotic membrane or autologous serum. PMID:20108493

Investigation of possible fs-LASIK induced retinal damage

NASA Astrophysics Data System (ADS)

Schumacher, S.; Sander, M.; Stolte, A.; Doepke, C.; Baumgaertner, W.; Lubatschowski, H.

2006-02-01

Rapid development of new laser technologies enabled the application of ultra short lasers in refractive surgery. Focused ultra short laser pulses in near-infrared spectral range can generate a laser induced breakdown (LIB) in the cornea, which will disrupt the tissue. Cutting depth and position can be established by varying the laser focus. The fs-LASIK technique allows both flap and lenticule to be formed by using fs-pulses without the presence of any mechanical impact. During the cutting process not all of the pulse energy is deposited into the cornea; approximately half of the remaining energy propagates through the eye and reaches the retina. Though defocused, the transmitted energy can still induce damage to the retina due to absorption by the retinal pigment epithelium and the transfer of thermal energy to surrounding tissue. The fs-LASIK process was simulated with two laser systems; one continous-wave and one in the fs-regime. For the simulation the exposure time and focusing numerical aperature which defines the retinal spot size were varied. The Damage thresholds of the laser beam exposed eyes were determined in terms of ophthalmoscopic and histopathologic observations.

Management of nontraumatic corneal perforation with tectonic drape patch and cyanoacrylate glue.

PubMed

Khalifa, Yousuf M; Bailony, M Rami; Bloomer, Michele M; Killingsworth, Daniel; Jeng, Bennie H

2010-10-01

To report a case of nontraumatic corneal perforation managed with a tectonic drape patch. Interventional case report. A 60-year-old patient with a corneal scar in his left eye likely secondary to herpes simplex virus interstitial keratitis underwent laser peripheral iridotomy for narrow angles. He developed progressive thinning of the cornea overlying the scar that led to a descemetocele and then ultimately a 1.2- × 1.7-mm perforation. Intraoperatively, several attempts were made to seal the perforation with cyanoacrylate glue, but the wound continued to leak. Sterile plastic drape that was on the surgical field was fashioned into a 2-mm-diameter patch, and the peripheral edge of the tectonic drape patch was glued over the perforation, successfully sealing the cornea. One week later, the drape patch was intact without leak, and a penetrating keratoplasty was carried out without complication. Tectonic drape patch technique for nontraumatic corneal perforations in which there is tissue loss is a viable temporizing option when cyanoacrylate glue alone fails and when there is no corneal tissue or amniotic membrane available to close the wound.

Fibrinogen, Riboflavin, and UVA to Immobilize a Corneal Flap—Conditions for Tissue Adhesion

PubMed Central

Littlechild, Stacy L.; Brummer, Gage; Zhang, Yuntao; Conrad, Gary W.

2012-01-01

Purpose. Laser-assisted in situ keratomileus (LASIK) creates a permanent flap that remains non-attached to the underlying laser-modified stroma. This lack of permanent adhesion is a liability. To immobilize a corneal flap, a protocol using fibrinogen (FIB), riboflavin (RF), and ultraviolet (UVA) light (FIB+RF+UVA) was devised to re-adhere the flap to the stroma. Methods. A model flap was created using rabbit (Oryctolagus cuniculus) and shark (Squalus acanthias) corneas. Solutions containing FIB and RF were applied between corneal strips as glue. Experimental corneas were irradiated with long wavelength (365 nm) UVA. To quantify adhesive strength between corneal strips, the glue-tissue interface was subjected to a constant force while a digital force gauge recorded peak tension. Results. In the presence of FIB, substantive non-covalent interactions occurred between rabbit corneal strips. Adhesiveness was augmented if RF and UVA also were applied, suggesting formation of covalent bonds. Additionally, exposing both sides of rabbit corneas to UVA generated more adhesion than exposure from one side, suggesting that RF in the FIB solution catalyzes formation of covalent bonds at only the interface between stromal molecules and FIB closest to the UVA. In contrast, in the presence of FIB, shark corneal strips interacted non-covalently more substantively than those of rabbits, and adhesion was not augmented by applying RF+UVA, from either or both sides. Residual RF could be rinsed away within 1 hour. Conclusions. Glue solution containing FIB and RF, together with UVA treatment, may aid immobilization of a corneal flap, potentially reducing risk of flap dislodgement. PMID:22589434

Salzmann's nodular degeneration of the cornea: a review and case series.

PubMed

Das, Sujata; Link, Barbara; Seitz, Berthold

2005-10-01

Salzmann's nodular degeneration is a rare, noninflammatory, slowly progressive, degenerative condition. Bluish-white nodules raised above the surface of the cornea characterize it. It has usually developed in corneas with a history of phlyctenulosis, trachoma, vernal keratoconjunctivitis, measles, scarlet fever, and various other viral diseases. However, today the majority of cases have been seen without recognized previous keratitis. It is composed of dense irregularly arranged collagen tissue with hyalinization between epithelium and Bowman's layer or beyond. Manual removal, phototherapeutic keratectomy (PTK) with or without the use of topical mitomycin-C, lamellar or penetrating keratoplasty have been used in the treatment of this disease. Salzmann's nodular degeneration does not seem to consist of one clinical entity. In some cases, elevated and pannus-like tissue can be separated easily from the corneal surface leaving Bowman's layer almost untouched. In these eyes, subsequent PTK may be necessary to smooth the surface. Recurrences are rare in these eyes. In contrast, some eyes (often with major peripheral vascularization) are left with deep defects in Bowman's layer and superficial stroma after difficult mechanical removal of nodules. In these eyes, multiple masking/laser ablation procedures are mandatory to acquire a homogenous surface. In our experience, the required laser ablation depth is significantly greater and the best-corrected visual acuity to be expected is reduced in contrast to the eyes with easy removal of the nodules. In these eyes recurrences seem to occur more frequently after treatment. Of 35 eyes documented to have Salzmann's nodular degeneration during the last 15 years in our department, 22 needed PTK treatment. Visual acuity increased from 0.4 to 0.7 on average. As a routine, laser ablation should be combined with previous conventional removal of nodules and excessive pannus tissue. By doing so, lamellar and penetrating keratoplasty techniques are hardly ever required in those eyes.

Comparative intraocular penetration of 4 fluoroquinolones after topical instillation.

PubMed

Chung, Jae Lim; Lim, Eun Hye; Song, Sang Wroul; Kim, Byung Yeop; Lee, Joon H; Mah, Francis S; Seo, Kyoung Yul

2013-07-01

To compare the intraocular penetration of 4 fluoroquinolone eye drops after topical instillation into rabbit eyes. The tested drugs were levofloxacin 1.5% (LVFX), gatifloxacin 0.3%, moxifloxacin 0.5% (MFLX), and besifloxacin 0.6% (BFLX). Forty-eight New Zealand white rabbits were randomly assigned into 2 groups. For group 1 (40 rabbits, 80 eyes), single instillation was performed, and tissue samples were acquired after 0.5, 1, 2, 4, and 6 hours. For group 2 (8 rabbits, 16 eyes), repeated instillation was performed (4 times, every 15 minutes), and tissues were acquired 1 hour after the fourth instillation. The drug concentrations in ocular tissues (cornea, aqueous, conjunctiva, and trisected vitreous) were analyzed with high-performance liquid chromatography. The AUC 0-6 h (area under the curve, in microgram.hour/gram) in group 1 and the mean concentration (in micrograms/gram) in group 2 for LVFX, gatifloxacin 0.3%, MFLX, and BFLX, respectively, were 22.97, 6.44, 13.54, and 3.29 and 22.60, 6.99, 13.69, and 1.91 in cornea; 5.66, 1.43, 3.38, and 0.42 and 5.52, 1.29, 2.47, and 0.19 in aqueous humor; 2.33, 0.91, 2.17, and 9.83 and 4.51, 0.78, 1.48, and 2.09 in bulbar conjunctiva; 0.243, 0.051, 0.134, and 0.018 and 0.182, 0.055, 0.122, and 0.015 in anterior vitreous; none of the drugs achieved enough concentration in equatorial and posterior vitreous. Repeated instillation resulted in approximately 2.1 times greater penetration than single instillation. LVFX and MFLX demonstrated good intraocular penetration particularly in cornea, aqueous humor, and anterior vitreous, and they may be considered the penetrative fluoroquinolones. BFLX showed high concentration in bulbar conjunctiva and may be considered the retentive fluoroquinolone.

Eye-bank preparation of endothelial tissue.

PubMed

Boynton, Grace E; Woodward, Maria A

2014-07-01

Eye-bank preparation of endothelial tissue for keratoplasty continues to evolve. Although eye-bank personnel have become comfortable and competent at Descemet's stripping automated endothelial keratoplasty (DSAEK), tissue preparation and tissue transport, optimization of preparation methods continues. Surgeons and eye-bank personnel should be up to date on the research in the field. As surgeons transit to Descemet's membrane endothelial keratoplasty (DMEK), eye banks have risen to the challenge of preparing tissue. Eye banks are refining their DMEK preparation and transport techniques. This article covers refinements to DSAEK tissue preparation, innovations to prepare DMEK tissue, and nuances to improve donor cornea tissue quality. As eye bank-supplied corneal tissue is the main source of tissue for many corneal surgeons, it is critical to stay informed about tissue handling and preparation. Ultimately, the surgeon is responsible for the transplantation, so involvement of clinicians in eye-banking practices and advocacy for pursuing meaningful research in this area will benefit clinical patient outcomes.

Tuna cornea as biomaterial for cardiac applications.

PubMed

Parravicini, Roberto; Cocconcelli, Flavio; Verona, Alessandro; Parravicini, Valeriano; Giuliani, Enrico; Barbieri, Alberto

2012-01-01

Among available biomaterials, cornea is almost completely devoid of cells and is composed only of collagen fibers oriented in an orderly pattern, which contributes to low antigenicity. Thunnus thynnus, the Atlantic bluefin tuna, is a fish with large eyes that can withstand pressures of approximately 10 MPa. We evaluated the potential of this tuna cornea in cardiac bioimplantation. Eyes from freshly caught Atlantic bluefin tuna were harvested and preserved in a fixative solution. Sterilized samples of corneal stroma were embedded in paraffin and stained with hematoxylin and eosin, and the histologic features were studied. Physical and mechanical resistance tests were performed in comparison with bovine pericardial strips and porcine mitral valves. Corneal material was implanted subcutaneously in 7 rats, to evaluate in vivo calcification rates. Mitral valves made from tuna corneal leaflets were implanted in 9 sheep. We found that the corneal tissue consisted only of parallel collagen fibers without evidence of vascular or neural structures. In tensile strength, the tuna corneal specimens were substantially similar to bovine pericardium. After 23 days, the rat-implanted samples showed no calcium or calcium salt deposition. Hydrodynamic and fatigue testing of valve prototypes yielded acceptable functional and long-term behavioral results. In the sheep, valvular performance was stable during the 180-day follow-up period, with no instrumental sign of calcification at the end of observation. We conclude that low antigenicity and favorable physical properties qualify tuna cornea as a potential material for durable bioimplantation. Further study is warranted.

Therapeutic penetrating keratoplasty in severe fungal keratitis using cryopreserved donor corneas

PubMed Central

Yao, Y-F; Zhang, Y-M; Zhou, P; Zhang, B; Qiu, W-Y; Tseng, S C G

2003-01-01

Aims: To investigate whether cryopreserved donor cornea could be used for therapeutic penetrating keratoplasty (PKP) to eradicate the infection, obviate complications, and preserve anatomical integrity in severe fungal keratitis. Methods: In this retrospective, consecutive case series, 45 eyes of 45 patients with severe fungal keratitis, which exhibited anterior chamber collapse, corneal perforation, and/or large suppurative corneal infiltrate, received therapeutic PKP after removal of the infected corneal tissue, irrigation of the anterior chamber by 0.2% fluconazole solution, iris dissection of fibrinoid membrane, and iridectomy and therapeutic PKP using corneas cryopreserved at −20°C. Results: Among 45 eyes, 39 eyes (86.7%) were successfully eradicated the fungal infection without recurrence and maintained their anatomical integrity without any complication. Four of 45 eyes (8.9%) showed postoperative rise of intraocular pressure, of which three were controlled with subsequent antiglaucoma surgeries, whereas one eye needed additional antiglaucoma medications. Two of 45 eyes (4.4%) were enucleated because of uncontrollable fungal infection and secondary retinal detachment, respectively. 23 eyes received subsequent optical PKP and, among them, 21 maintained clear corneal grafts and two suffered from graft failure due to allograft rejections. Conclusion: Cryopreserved donor corneas are effective substitutes in therapeutic PKP to control severe fungal corneal infection and preserve the global integrity, and may offer additional advantages over conventional PKP in reducing allograft rejection, eradicating fungal infection during the postoperative period, and improving the success of optical PKP for visual rehabilitation. PMID:12714387

Risk factors for donor endothelial loss in eye bank-prepared posterior lamellar corneal tissue for descemet stripping automated endothelial keratoplasty.

PubMed

Liu, Yu-Chi; Alvarez Paraz, Carisa M; Cajucom-Uy, Howard Yu; Agahari, Djoni; Sethuraman, Selvam; Tan, Donald T-H; Mehta, Jodhbir S

2014-07-01

The aim of this study was to investigate donor, tissue, and precut procedure risk factors for endothelial cell density (ECD) loss in posterior lamellar corneal tissue preparation by an eye bank for Descemet stripping automated endothelial keratoplasty. A total of 259 corneoscleral rims precut by the Singapore Eye Bank from October 2011 to August 2013 were evaluated. Donor characteristics, tissue characteristics, and precut procedure parameters were analyzed. The mean donor age was 57.18 ± 11.35 years, and the mean cutting transition time was 4.16 ± 0.75 seconds. The mean ECD was 2826 ± 225 and 2787 ± 224 cells per square millimeter before and after precutting, respectively, with an average ECD change of -1.38% ± 3.28%. The precutting procedure failure rate was 1.2%. Mutivariate regression analysis showed that an older donor age, a higher ECD before cutting, and a slower cutting transition speed were significant factors. Corneas with an ECD >2800 cells per square millimeter before precutting, cutting transition time >5.5 seconds, and corneas with donor age >65 years were significantly more likely to have greater than 5% ECD loss after precutting (odds ratio, 6.42, 1.66, and 1.62; 95% confidence interval, 1.44-29.43, 1.45-2.72, and 1.66-5.82, respectively). Donor source, death-to-preservation time (range, 0.67-10.88 hours), death-to-precutting time (range, 0-7 days), and graft thickness (range, 43-232 μm) were not statistically significant factors. The ECD loss in the precut tissue prepared by the eye bank was very low. The risk factors identified provide better understanding of how to improve the quality and safety profiles when preparing graft tissue for Descemet stripping automated endothelial keratoplasty.

Thermal and infrared-diode laser effects on indocyanine-green-treated corneal collagen

NASA Astrophysics Data System (ADS)

Timberlake, George T.; Patmore, Ann; Shallal, Assaad; McHugh, Dominic; Marshall, John

1993-07-01

It has been suggested that laser welds of collagenous tissues form by interdigitation and chemical bonding of thermally 'unraveled' collagen fibrils. We investigated this proposal by attempting to weld highly collagenous, avascular corneal tissue with an infrared (IR) diode laser as follows. First, the temperature at which corneal collagen shrinks and collagen fibrils 'split' into subfibrillary components was determined. Second, since use of a near-IR laser wavelength necessitated addition of an absorbing dye (indocyanine green (ICG) to the cornea, we measured absorption spectra of ICG-treated tissue to ensure that peak ICG absorbance did not change markedly when ICG was present in the cornea. Third, using gel electrophoresis of thermally altered corneal collagen, we searched for covalently crosslinked compounds predicted by the proposed welding mechanism. Finally, we attempted to weld partial thickness corneal incisions infused with ICG. Principal experimental findings were as follows: (1) Human corneal (type I) collagen splits into subfibrillary components at approximately 63 degree(s)C, the same temperature that produces collagen shrinkage. (2) Peak ICG absorption does not change significantly in corneal stroma or with laser heating. (3) No evidence was found for the formation of novel compounds or the loss of proteins as a result of tissue heating. All tissue treated with ICG, however, exhibited a novel 244 kD protein band indicating chemical activity between collagen and corneal stromal components. (4) Laser welding corneal incisions was unsuccessful possibly due to shrinkage of the sides of the incision, lack of incision compression during heating, or a less than optimal combination of ICG concentration and radiant exposure. In summary, these experiments demonstrate the biochemical and morphological complexity of ICG-enhanced IR laser-tissue welding and the need for further investigation of laser welding mechanisms.

Expression and localization of epithelial stem cell and differentiation markers in equine skin, eye and hoof.

PubMed

Linardi, Renata L; Megee, Susan O; Mainardi, Sarah R; Senoo, Makoto; Galantino-Homer, Hannah L

2015-08-01

The limited characterization of equine skin, eye and hoof epithelial stem cell (ESC) and differentiation markers impedes the investigation of the physiology and pathophysiology of these tissues. To characterize ESC and differentiation marker expression in epithelial tissues of the equine eye, haired skin and hoof capsule. Indirect immunofluorescence microscopy and immunoblotting were used to detect expression and tissue localization of keratin (K) isoforms K3, K10, K14 and K124, the transcription factor p63 (a marker of ESCs) and phosphorylated p63 [pp63; a marker of ESC transition to transit-amplifying (TA) cell] in epithelial tissues of the foot (haired skin, hoof coronet and hoof lamellae) and the eye (limbus and cornea). Expression of K14 was restricted to the basal layer of epidermal lamellae and to basal and adjacent suprabasal layers of the haired skin, coronet and corneal limbus. Coronary and lamellar epidermis was negative for both K3 and K10, which were expressed in the cornea/limbus epithelium and haired skin epidermis, respectively. Variable expression of p63 with relatively low to high levels of phosphorylation was detected in individual basal and suprabasal cells of all epithelial tissues examined. To the best of the author's knowledge, this is the first report of the characterization of tissue-specific keratin marker expression and the localization of putative epithelial progenitor cell populations, including ESCs (high p63 expression with low pp63 levels) and TA cells (high expression of both p63 and pp63), in the horse. These results will aid further investigation of epidermal and corneal epithelial biology and regenerative therapies in horses. © 2015 ESVD and ACVD.

[Sutureless lamellar keratoplasty by microkeratome combined with fibrin tissue adhesive in rabbits].

PubMed

Chen, Wei; Qu, Jia; Lu, Fan; Zhu, Ren-yuan

2004-05-01

To evaluate the feasibility and safety of sutureless lamellar keratoplasty by microkeratome combined with fibrin tissue adhesive. Twenty-four New Zealand white rabbits were divided into two groups, the donor grafts and recipient beds were made by the microkeratome, the grafts were glued over the stoma bed using the commercial product Tisseel in one group; and grafts without tissue adhesive were used as the control group. Corneal refractive power was measured by automated keratometer preoperatively and in 3 days, 2 weeks, 1 and 3 months postoperatively. Rejection and cornea transparency were observed. Confocal microscopy was used to observe corneal wound healing response and to measure the keratocyte and endothelium densities in vivo. Corneal wound healing was also evaluated using light and fluorescence microscopy. Ninety-two percent (11/12 eyes) of the glued grafts were retained in the Tisseel group, whereas all grafts were lost in the control group. All survived grafts were clear 1 month after surgery. However, in the control group, severe haze in the grafts occurred 2 weeks postoperatively. Confocal microscopy showed that there was a significant decrease of the keratocyte density surrounding the lenticule-host interface, and no changes occurred in the posterior keratocyte and endothelium. Histopathologic observations demonstrated the presence of a line of amorphous eosinophilic substance in the lenticule-host interface at 3 days after surgery, but the line disappeared after 1 month. Fluorescence microscopy showed no detectable regenerated stromal tissue. This initial study demonstrates sutureless optical lamellar keratoplasty performed by microkeratome combined with fibrin tissue adhesive is a simple and safe technique. Stromal wound healing response to this surgery is minimal. Fibrin tissue adhesive has no influence on the cornea optical property.

Experimental study on the healing process following laser welding of the cornea.

PubMed

Rossi, Francesca; Pini, Roberto; Menabuoni, Luca; Mencucci, Rita; Menchini, Ugo; Ambrosini, Stefano; Vannelli, Gabriella

2005-01-01

An experimental study evaluating the application of laser welding of the cornea and the subsequent healing process is presented. The welding of corneal wounds is achieved after staining the cut walls with a solution of the chromophore indocyanine green, and irradiating them with a diode laser (810 nm) operating at low power (60 to 90 mW). The result is a localized heating of the cut, inducing controlled welding of the stromal collagen. In order to optimize this technique and to study the healing process, experimental tests, simulating cataract surgery and penetrating keratoplasty, were performed on rabbits: conventional and laser-induced suturing of corneal wounds were thus compared. A follow-up study 7 to 90 days after surgery was carried out by means of objective and histological examinations, in order to optimize the welding technique and to investigate the subsequent healing process. The analyses of the laser-welded corneas evidenced a faster and more effective restoration of the architecture of the stroma. No thermal damage of the welded stroma was detected, nor were there foreign body reactions or other inflammatory processes. Copyright 2005 Society of Photo-Optical Instrumentation Engineers.

Laser tissue welding in ophthalmic surgery.

PubMed

Rossi, Francesca; Matteini, Paolo; Ratto, Fulvio; Menabuoni, Luca; Lenzetti, Ivo; Pini, Roberto

2008-09-01

Laser welding of ocular tissues is an alternative technique or adjunct to conventional suturing in ophthalmic surgery. It is based on the photothermal interaction of laser light with the main components of the extracellular matrix of connective tissues. The advantages of the welding procedure with respect to standard suturing and stapling are reduced operation times, lesser inflammation, faster healing and increased ability to induce tissue regeneration. The procedure we set up is based on the use of an infrared diode laser in association with the topical application of the chromophore Indocyanine Green. Laser light may be delivered either continuously or in pulses, thus identifying two different techniques that have been applied clinically in various types of transplants of the cornea.

Eye drop delivery of nano-polymeric micelle formulated genes with cornea-specific promoters.

PubMed

Tong, Yaw-Chong; Chang, Shwu-Fen; Liu, Chia-Yang; Kao, Winston W-Y; Huang, Chong Heng; Liaw, Jiahorng

2007-11-01

This study evaluates the eye drop delivery of genes with cornea-specific promoters, i.e., keratin 12 (K12) and keratocan (Kera3.2) promoters, by non-ionic poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) (PEO-PPO-PEO) polymeric micelles (PM) to mouse and rabbit eyes, and investigates the underlying mechanisms. Three PM-formulated plasmids (pCMV-Lac Z, pK12-Lac Z and pKera3.2-Lac Z) containing the Lac Z gene for beta-galactosidase (beta-Gal) whose expression was driven by the promoter of either the cytomegalovirus early gene, the keratin 12 gene or the keratocan gene, were characterized by critical micelle concentration (CMC), dynamic light scattering (DLS), and atomic force microscopy (AFM). Transgene expression in ocular tissue after gene delivery was analyzed by 5-bromo-4-chloro-3-indolyl-beta-D-galactoside (X-Gal) color staining, 1,2-dioxetane beta-Gal enzymatic activity measurement, and real-time polymerase chain reaction (PCR) analysis. The delivery mechanisms of plasmid-PM on mouse and rabbit corneas were evaluated by EDTA and RGD (arginine-glycine-aspartic acid) peptide. The sizes of the three plasmid-PM complexes were around 150-200 nm with unimodal distribution. Enhanced stability was found for three plasmid-PM formulations after DNase I treatment. After six doses of eye drop delivery of pK12-Lac Z-PM three times a day, beta-Gal activity was significantly increased in both mouse and rabbit corneas. Stroma-specific Lac Z expression was only found in pKera3.2-Lac Z-PM-treated animals with pretreatment by 5 mM EDTA, an opener of junctions. Lac Z gene expression in both pK12-Lac Z-PM and pKera3.2-Lac Z-PM delivery groups was decreased by RGD peptide pretreatment. Cornea epithelium- and stroma-specific gene expression could be achieved using cornea-specific promoters of keratin 12 and keratocan genes, and the gene was delivered with PM formulation through non-invasive, eye drop in mice and rabbits. The transfection mechanism of plasmid-PM may involve endocytosis and particle size dependent paracellular transport. 2007 John Wiley & Sons, Ltd

Global Survey of Corneal Transplantation and Eye Banking.

PubMed

Gain, Philippe; Jullienne, Rémy; He, Zhiguo; Aldossary, Mansour; Acquart, Sophie; Cognasse, Fabrice; Thuret, Gilles

2016-02-01

Corneal transplantation restores visual function when visual impairment caused by a corneal disease becomes too severe. It is considered the world's most frequent type of transplantation, but, to our knowledge, there are no exhaustive data allowing measurement of supply and demand, although such data are essential in defining local, national, and global strategies to fight corneal blindness. To describe the worldwide situation of corneal transplantation supply and demand. Data were collected between August 2012 and August 2013 from a systematic review of published literature in parallel with national and international reports on corneal transplantation and eye banking. In a second step, eye bank staff and/or corneal surgeons were interviewed on their local activities. Interviews were performed during international ophthalmology or eye-banking congresses or by telephone or email. Countries' national supply/demand status was classified using a 7-grade system. Data were collected from 148 countries. Corneal transplantation and corneal procurements per capita in each country. In 2012, we identified 184,576 corneal transplants performed in 116 countries. These were procured from 283,530 corneas and stored in 742 eye banks. The top indications were Fuchs dystrophy (39% of all corneal transplants performed), a primary corneal edema mostly affecting elderly individuals; keratoconus (27%), a corneal disease that slowly deforms the cornea in young people; and sequellae of infectious keratitis (20%). The United States, with 199.10-6 corneal transplants per capita, had the highest transplantation rate, followed by Lebanon (122.10-6) and Canada (117.10-6), while the median of the 116 transplanting countries was 19.10-6. Corneas were procured in only 82 countries. Only the United States and Sri Lanka exported large numbers of donor corneas. About 53% of the world's population had no access to corneal transplantation. Our survey globally quantified the considerable shortage of corneal graft tissue, with only 1 cornea available for 70 needed. Efforts to encourage cornea donation must continue in all countries, but it is also essential to develop alternative and/or complementary solutions, such as corneal bioengineering.

Controlling human corneal stromal stem cell contraction to mediate rapid cell and matrix organization of real architecture for 3-dimensional tissue equivalents.

PubMed

Mukhey, Dev; Phillips, James B; Daniels, Julie T; Kureshi, Alvena K

2018-02-01

The architecture of the human corneal stroma consists of a highly organized extracellular matrix (ECM) interspersed with keratocytes. Their progenitor cells; corneal stromal stem cells (CSSC) are located at the periphery, in the limbal stroma. A highly organized corneal ECM is critical for effective transmission of light but this structure may be compromised during injury or disease, resulting in loss of vision. Re-creating normal organization in engineered tissue equivalents for transplantation often involves lengthy culture times that are inappropriate for clinical use or utilisation of synthetic substrates that bring complications such as corneal melting. CSSC have great therapeutic potential owing to their ability to reorganize a disorganized matrix, restoring transparency in scarred corneas. We examined CSSC contractile behavior to assess whether this property could be exploited to rapidly generate cell and ECM organization in Real Architecture For 3D Tissues (RAFT) tissue equivalents (TE) for transplantation. Free-floating collagen gels were characterized to assess contractile behavior of CSSC and establish optimum cell density and culture times. To mediate cell and collagen organization, tethered collagen gels seeded with CSSC were cultured and subsequently stabilized with the RAFT process. We demonstrated rapid creation of biomimetic RAFT TE with tunable structural properties. These displayed three distinct regions of varying degrees of cellular and collagen organization. Interestingly, increased organization coincided with a dramatic loss of PAX6 expression in CSSC, indicating rapid differentiation into keratocytes. The organized RAFT TE system could be a useful bioengineering tool to rapidly create an organized ECM while simultaneously controlling cell phenotype. For the first time, we have demonstrated that human CSSC exhibit the phenomenon of cellular self-alignment in tethered collagen gels. We found this mediated rapid co-alignment of collagen fibrils and thus subsequently exploited this property in vitro to improve the architecture of engineered RAFT tissue equivalents of the corneal stroma. Existing techniques are extremely lengthy and carry significant risk and cost for GMP manufacture. This rapid and tunable technique takes just 8 h of culture and is therefore ideal for clinical manufacture, creating biomimetic tissue equivalents with both cellular and ECM organization. Thus, cellular self-alignment can be a useful bioengineering tool for the development of organized tissue equivalents in a variety of applications. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Spectral optical coherence tomography: a novel technique for cornea imaging.

PubMed

Kaluzny, Bartłomiej J; Kaluzy, Bartłomiej J; Kałuzny, Jakub J; Szkulmowska, Anna; Gorczyńska, Iwona; Szkulmowski, Maciej; Bajraszewski, Tomasz; Wojtkowski, Maciej; Targowski, Piotr

2006-09-01

Spectral optical coherence tomography (SOCT) is a new, noninvasive, noncontact, high-resolution technique that provides cross-sectional images of the objects that weakly absorb and scatter light. SOCT, because of very short acquisition time and high sensitivity, is capable of providing tomograms of substantially better quality than the conventional OCT. The aim of this paper is to show the application of the SOCT to cross-sectional imaging of the cornea and its pathologies. Eleven eyes with different corneal pathologies were examined with a slit lamp and the use of a prototype SOCT instrument constructed in the Institute of Physics, Nicolaus Copernicus University, Toruń, Poland. Our SOCT system provides high-resolution (4 microm axial, 10 microm transversal) tomograms composed of 3000-5000 A-scans with an acquisition time of 120-200 ms. The quality of the images is adequate for detailed cross-sectional evaluation of various corneal pathologies. Objective assessment of the localization, size, shape, and light-scattering properties of the changed tissue is possible. Corneal and epithelial thickness and the depth and width of lesions can be estimated. SOCT technique allows acquiring clinically valuable cross-sectional optical biopsy of the cornea and its pathologies.

Corneal cut closure using temperature-controlled CO2 laser soldering system.

PubMed

Tal, Kfir; Strassmann, Eyal; Loya, Nino; Ravid, Avi; Kariv, Noam; Weinberger, Dov; Katzir, Abraham; Gaton, Dan D

2015-05-01

We aimed to evaluate the effectiveness of temperature-controlled laser soldering for repair of large perforated corneas in a porcine model. Eight Yorkshire pigs aged 6 months underwent 6-mm-deep 180° crescent-shaped trephination of the central corneas. Right corneal injuries were repaired by placement of 47 % bovine albumin along the cut followed by CO2 laser soldering (power density 16 W/cm(2)) to a target temperature of 65(°). Left corneal injuries were repaired with 10/0 nylon sutures. The groups were compared for operative time, leakage, and histopathological findings. Mean tissue temperature was 63 ± 4 °C. Mean operative time was 31.57 ± 2.8 min in laser-soldered eyes and 41.38 ± 2.3 min in controls (p < 0.0001, unpaired Student's t test). Compared to controls, the soldered corneas had less neovascularization, complete re-epithelization, and mild stromal inflammation. There was no leakage in either group. Combined CO2 laser and radiometer is effective for the in vivo repair of corneal cuts. These results have important implications for modern corneal surgery. Further studies are needed in the clinical setting.

Novel Lutein Loaded Lipid Nanoparticles on Porcine Corneal Distribution

PubMed Central

Liu, Chi-Hsien; Chiu, Hao-Che; Wu, Wei-Chi; Sahoo, Soubhagya Laxmi; Hsu, Ching-Yun

2014-01-01

Topical delivery has the advantages including being user friendly and cost effective. Development of topical delivery carriers for lutein is becoming an important issue for the ocular drug delivery. Quantification of the partition coefficient of drug in the ocular tissue is the first step for the evaluation of delivery efficacy. The objectives of this study were to evaluate the effects of lipid nanoparticles and cyclodextrin (CD) on the corneal lutein accumulation and to measure the partition coefficients in the porcine cornea. Lipid nanoparticles combined with 2% HPβCD could enhance lutein accumulation up to 209.2 ± 18 (μg/g) which is 4.9-fold higher than that of the nanoparticles. CD combined nanoparticles have 68% of drug loading efficiency and lower cytotoxicity in the bovine cornea cells. From the confocal images, this improvement is due to the increased partitioning of lutein to the corneal epithelium by CD in the lipid nanoparticles. The novel lipid nanoparticles could not only improve the stability and entrapment efficacy of lutein but also enhance the lutein accumulation and partition in the cornea. Additionally the corneal accumulation of lutein was further enhanced by increasing the lutein payload in the vehicles. PMID:25101172

Lasers in ophthalmology

NASA Astrophysics Data System (ADS)

Mardin, Christian Y.; Tornow, Ralf P.; Kruse, Friedrich E.

Lasers are widely used in Ophthalmology. The unique transparent tissues of the eye's cornea, lens and vitreous body make lasers an ideal medium for diagnosis and therapy. They are incorporated in well established commercially available ophthalmic instruments but they also play an important role in new developments and research work. This paper gives a short overview on selected laser applications in our department.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ionin, Andrey A.; Kozhushko, Svetlana E.; Kudryashov, Sergey I.

We have successfully fabricated in vitro femtosecond laser micro-incisions inside cornea and--for the first time--inside sclera mildly pre-cleared by a biocompatible and clinically safe (non-toxic) natural agent (replacive refractive index-matching 40%-glucose solution in water), with the tissues taken as fresh cool cuts of human cadaver eyes, and reported on basic operational conditions of the micro-surgical procedures.

Screening of post-mortem tissue donors for Coxiella burnetii infection after large outbreaks of Q fever in The Netherlands

PubMed Central

2014-01-01

Background After the largest outbreaks of Q fever ever recorded in history occurred in the Netherlands, concern arose that Coxiella may be transmitted via donated tissues of latent or chronically infected donors. The Dutch Health Council recently advised to screen tissue donors, donating high risk tissues, for Coxiella infection. Methods After validation of an enzyme immunoassay (EIA) test for IgG antibodies against phase 2 of C. burnetii for use on post-mortem samples, serum samples of 1033 consecutive Dutch post-mortem tissue donors were tested for IgG antibodies against phase 2 of C. burnetii. Confirmation of reactive results was done by immunofluorescence assay (IFA). All available tissues (corneas, heart valves, skin and bone marrow) from donors with IgG reactivity were tested for presence of Coxiella DNA by PCR. Risk factors for IgG reactivity were investigated. Results After validation of the tests for use on post-mortem samples, 50/1033 donors (4.8%) screened positive for phase 2 anti-Coxiella IgG by EIA, and 31 were confirmed by IFA (3.0%). One donor showed a serological profile compatible with chronic infection. All tested tissues (25 corneas, 6 heart valves, 4 skin and 3 bone marrow) from donors with IgG reactivity tested negative for the presence of Coxiella DNA. Except for living in a postal code area with a high number of Q fever notifications, no risk factors for IgG reactivity were found. Conclusions The strong correlation between notifications and seroprevalence confirms that the used assays are sufficiently specific for use on post-mortem samples, although one has to be aware of differences between batches. Thus, this study provides a validated method for screening tissue donors for infection with Coxiella burnetii that can be used in future outbreaks. PMID:24393298

[Establishment of goat limbal stem cell strain expressing Venus fluorescent protein and construction of limbal epithelial sheets].

PubMed

Yin, Jiqing; Liu, Wenqiang; Liu, Chao; Zhao, Guimin; Zhang, Yihua; Liu, Weishuai; Hua, Jinlian; Dou, Zhongying; Lei, Anmin

2010-12-01

The integrity and transparency of cornea plays a key role in vision. Limbal Stem Cells (LSCs) are precursors of cornea, which are responsible for self-renewal and replenishing corneal epithelium. Though it is successful to cell replacement therapy for impairing ocular surface by Limbal Stem Cell Transplantation (LSCT), the mechanism of renew is unclear after LSCT. To real time follow-up the migration and differentiation of corneal transplanted epithelial cells after transplanting, we transfected venus (a fluorescent protein gene) into goat LSCs, selected with G418 and established a stable transfected cell line, named GLSC-V. These cells showed green fluorescence, and which could maintain for at least 3 months. GLSC-V also were positive for anti-P63 and anti-Integrinbeta1 antibody by immunofluorescent staining. We founded neither GLSC-V nor GLSCs expressed keratin3 (k3) and keratinl2 (k12). However, GLSC-V had higher levels in expression of p63, pcna and venus compared with GLSCs. Further, we cultivated the cells on denude amniotic membrane to construct tissue engineered fluorescent corneal epithelial sheets. Histology and HE staining showed that the constructed fluorescent corneal epithelial sheets consisted of 5-6 layers of epithelium. Only the lowest basal cells of fluorescent corneal epithelial sheets expressed P63 analyzed by immunofluorescence, but not superficial epithelial cells. These results showed that our constructed fluorescent corneal epithelial sheets were similar to the normal corneal epithelium in structure and morphology. This demonstrated that they could be transplanted for patents with corneal impair, also may provide a foundation for the study on the mechanisms of corneal epithelial cell regeneration after LSCT.

Transporter targeted gatifloxacin prodrugs: synthesis, permeability, and topical ocular delivery.

PubMed

Vooturi, Sunil K; Kadam, Rajendra S; Kompella, Uday B

2012-11-05

In this work, we aim to design and synthesize prodrugs of gatifloxacin targeting organic cation transporter (OCT), monocarboxylate transporter (MCT), and ATB (0, +) transporters and to identify a prodrug with enhanced delivery to the back of the eye. Dimethylamino-propyl, carboxy-propyl, and amino-propyl(2-methyl) derivatives of gatifloxacin (GFX), DMAP-GFX, CP-GFX, and APM-GFX, were designed and synthesized to target OCT, MCT, and ATB (0, +) transporters, respectively. An LC-MS method was developed to analyze drug and prodrug levels in various studies. Solubility and log D (pH 7.4) were measured for prodrugs and the parent drug. The permeability of the prodrugs was determined in the cornea, conjunctiva, and sclera-choroid-retinal pigment epitheluim (SCRPE) and compared with gatifloxacin using an Ussing chamber assembly. Permeability mechanisms were elucidated by determining the transport in the presence of transporter specific inhibitors. 1-Methyl-4-phenylpyridinium iodide (MPP+), nicotinic acid sodium salt, and α-methyl-DL-tryptophan were used to inhibit OCT, MCT, and ATB (0, +) transporters, respectively. A prodrug selected based on in vitro studies was administered as an eye drop to pigmented rabbits, and the delivery to various eye tissues including vitreous humor was compared with gatifloxacin dosing. DMAP-GFX exhibited 12.8-fold greater solubility than GFX. All prodrugs were more lipophilic, with the measured log D (pH 7.4) values ranging from 0.05 to 1.04, when compared to GFX (log D: -1.15). DMAP-GFX showed 1.4-, 1.8-, and 1.9-fold improvement in permeability across the cornea, conjunctiva, and SCRPE when compared to GFX. Moreover, it exhibited reduced permeability in the presence of MPP+ (competitive inhibitor of OCT), indicating OCT-mediated transport. CP-GFX showed 1.2-, 2.3-, and 2.5-fold improvement in permeability across the cornea, conjunctiva, and SCRPE, respectively. In the presence of nicotinic acid (competitive inhibitor of MCT), the permeability of CP-GFX was reduced across the conjunctiva. However, the cornea and SCRPE permeability of CP-GFX was not affected by nicotinic acid. APM-GFX did not show any improvement in permeability when compared to GFX across the cornea, conjunctiva, and SCRPE. Based on solubility and permeability, DMAP-GFX was selected for in vivo studies. DMAP-GFX showed 3.6- and 1.95-fold higher levels in vitreous humor and CRPE compared to that of GFX at 1 h after topical dosing. In vivo conversion of DMAP-GFX prodrug to GFX was quantified in tissues isolated at 1 h after dosing. The parent drug-to-prodrug ratio was 8, 70, 24, 21, 29, 13, 55, and 60% in the cornea, conjunctiva, iris-ciliary body, aqueous humor, sclera, CRPE, retina, and vitreous humor, respectively. In conclusion, DMAP-GFX prodrug enhanced solubility, log D, as well as OCT mediated delivery of gatifloxacin to the back of the eye.

Depth-resolved monitoring of analytes diffusion in ocular tissues

NASA Astrophysics Data System (ADS)

Larin, Kirill V.; Ghosn, Mohamad G.; Tuchin, Valery V.

2007-02-01

Optical coherence tomography (OCT) is a noninvasive imaging technique with high in-depth resolution. We employed OCT technique for monitoring and quantification of analyte and drug diffusion in cornea and sclera of rabbit eyes in vitro. Different analytes and drugs such as metronidazole, dexamethasone, ciprofloxacin, mannitol, and glucose solution were studied and whose permeability coefficients were calculated. Drug diffusion monitoring was performed as a function of time and as a function of depth. Obtained results suggest that OCT technique might be used for analyte diffusion studies in connective and epithelial tissues.

Precut cornea for Descemet's stripping endothelial keratoplasty: experience at a single eye bank.

PubMed

Mohamed, Ashik; Chaurasia, Sunita; Chandragiri, Venkataswamy; Kandhibanda, Srinivas; Gunnam, Srinivas; Garg, Prashant

2017-06-01

The aim of the study is to describe the experience with precut facility for endothelial keratoplasty at a single eye bank affiliated to a tertiary eye care center in India. Data on precut tissues from Nov 2012 to Dec 2014 were retrospectively reviewed from the electronic database of the eye bank of a tertiary eye care center in South India. Donor characteristic data including donor age, precut and postcut endothelial cell density (ECD), recipient age, and thickness of graft were collected. The number of precuts increased from 42 in 2012 (Nov and Dec) and 422 in 2013 to 584 in 2014. Of the total of 1048 precuts, seven (0.67 %) were miscut and could not be utilized for transplants. The donor age ranged from 2 to 89 years. A mean change of 43.6 ± 325.2 cells/mm 2 in ECD was noted after cut, proportional increase in mean being 1.9 %. The change in ECD after cut was negatively correlated with ECD before cut. The recipient age ranged from 1 to 89 years. The median thickness of donor lenticule after cut was 148 µm (interquartile range 131-166 µm). Analysis of precut donor corneas from a single eye bank shows that the ECD of the processed tissues was excellent for transplantation. The tissue wastage in the hands of eye bank personal was minimal.

Non-contact rapid optical coherence elastography by high-speed 4D imaging of elastic waves

NASA Astrophysics Data System (ADS)

Song, Shaozhen; Yoon, Soon Joon; Ambroziński, Łukasz; Pelivanov, Ivan; Li, David; Gao, Liang; Shen, Tueng T.; O'Donnell, Matthew; Wang, Ruikang K.

2017-02-01

Shear wave OCE (SW-OCE) uses an OCT system to track propagating mechanical waves, providing the information needed to map the elasticity of the target sample. In this study we demonstrate high speed, 4D imaging to capture transient mechanical wave propagation. Using a high-speed Fourier domain mode-locked (FDML) swept-source OCT (SS-OCT) system operating at 1.62 MHz A-line rate, the equivalent volume rate of mechanical wave imaging is 16 kvps (kilo-volumes per second), and total imaging time for a 6 x 6 x 3 mm volume is only 0.32 s. With a displacement sensitivity of 10 nanometers, the proposed 4D imaging technique provides sufficient temporal and spatial resolution for real-time optical coherence elastography (OCE). Combined with a new air-coupled, high-frequency focused ultrasound stimulator requiring no contact or coupling media, this near real-time system can provide quantitative information on localized viscoelastic properties. SW-OCE measurements are demonstrated on tissue-mimicking phantoms and porcine cornea under various intra-ocular pressures. In addition, elasticity anisotropy in the cornea is observed. Images of the mechanical wave group velocity, which correlates with tissue elasticity, show velocities ranging from 4-20 m/s depending on pressure and propagation direction. These initial results strong suggest that 4D imaging for real-time OCE may enable high-resolution quantitative mapping of tissue biomechanical properties in clinical applications.

Full-field OCT: applications in ophthalmology

NASA Astrophysics Data System (ADS)

Grieve, Kate; Dubois, Arnaud; Paques, Michel; Le Gargasson, Jean-Francois; Boccara, Albert C.

2005-04-01

We present images of ocular tissues obtained using ultrahigh resolution full-field OCT. The experimental setup is based on the Linnik interferometer, illuminated by a tungsten halogen lamp. En face tomographic images are obtained in real-time without scanning by computing the difference of two phase-opposed interferometric images recorded by a high-resolution CCD camera. A spatial resolution of 0.7 μm × 0.9 μm (axial × transverse) is achieved thanks to the short source coherence length and the use of high numerical aperture microscope objectives. A detection sensitivity of 90 dB is obtained by means of image averaging and pixel binning. Whole unfixed eyes and unstained tissue samples (cornea, lens, retina, choroid and sclera) of ex vivo rat, mouse, rabbit and porcine ocular tissues were examined. The unprecedented resolution of our instrument allows cellular-level resolution in the cornea and retina, and visualization of individual fibers in the lens. Transcorneal lens imaging was possible in all animals, and in albino animals, transscleral retinal imaging was achieved. We also introduce our rapid acquisition full-field optical coherence tomography system designed to accommodate in vivo ophthalmologic imaging. The variations on the original system technology include the introduction of a xenon arc lamp as source, and rapid image acquisition performed by a high-speed CMOS camera, reducing acquisition time to 5 ms per frame.

Noncontact phase-sensitive dynamic optical coherence elastography at megahertz rate

NASA Astrophysics Data System (ADS)

Singh, Manmohan; Wu, Chen; Liu, Chih-Hao; Li, Jiasong; Schill, Alexander; Nair, Achuth; Kistenev, Yury V.; Larin, Kirill V.

2016-03-01

Dynamic optical coherence elastography (OCE) techniques have shown great promise at quantitatively obtaining the biomechanical properties of tissue. However, the majority of these techniques have required multiple temporal OCT acquisitions (M-B mode) and corresponding excitations, which lead to clinically unfeasible acquisition times and potential tissue damage. Furthermore, the large data sets and extended laser exposures hinder their translation to the clinic, where patient discomfort and safety are critical criteria. In this work we demonstrate noncontact true kilohertz frame-rate dynamic optical coherence elastography by directly imaging a focused air-pulse induced elastic wave with a home-built phase-sensitive OCE system based on a 4X buffered Fourier Domain Mode Locked swept source laser with an A-scan rate of ~1.5 MHz. The elastic wave was imaged at a frame rate of ~7.3 kHz using only a single excitation. In contrast to previous techniques, successive B-scans were acquired over the measurement region (B-M mode) in this work. The feasibility of this method was validated by quantifying the elasticity of tissue-mimicking agar phantoms as well as porcine corneas ex vivo at different intraocular pressures. The results demonstrate that this method can acquire a depth-resolved elastogram in milliseconds. The reduced data set enabled a rapid elasticity assessment, and the ultra-fast acquisition speed allowed for a clinically safe laser exposure to the cornea.

Colony Stimulating Factor-1 Receptor Expressing Cells Infiltrating the Cornea Control Corneal Nerve Degeneration in Response to HSV-1 Infection

PubMed Central

Chucair-Elliott, Ana J.; Gurung, Hem R.; Carr, Meghan M.; Carr, Daniel J. J.

2017-01-01

Purpose Herpes simplex virus type-1 (HSV-1) is a leading cause of neurotrophic keratitis, characterized by decreased or absent corneal sensation due to damage to the sensory corneal innervation. We previously reported the elicited immune response to infection contributes to the mechanism of corneal nerve regression/damage during acute HSV-1 infection. Our aim is to further establish the involvement of infiltrated macrophages in the mechanism of nerve loss upon infection. Methods Macrophage Fas-Induced Apoptosis (MAFIA) transgenic C57BL/6 mice were systemically treated with AP20187 dimerizer or vehicle (VEH), and their corneas, lymph nodes, and blood were assessed for CD45+CD11b+GFP+ cell depletion by flow cytometry (FC). Mice were ocularly infected with HSV-1 or left uninfected. At 2, 4, and/or 6 days post infection (PI), corneas were assessed for sensitivity and harvested for FC, nerve structure by immunohistochemistry, viral content by plaque assay, soluble factor content by suspension array, and activation of signaling pathways by Western blot analysis. C57BL6 mice were used to compare to the MAFIA mouse model. Results MAFIA mice treated with AP20187 had efficient depletion of CD45+CD11b+GFP+ cells in the tissues analyzed. The reduction of CD45+CD11b+GFP+ cells recruited to the infected corneas of AP20187-treated mice correlated with preservation of corneal nerve structure and function, decreased protein concentration of inflammatory cytokines, and decreased STAT3 activation despite no changes in viral content in the cornea compared to VEH-treated animals. Conclusions Our results suggest infiltrated macrophages are early effectors in the nerve regression following HSV-1 infection. We propose the neurodegeneration mechanism involves macrophages, local up-regulation of IL-6, and activation of STAT3. PMID:28903153

Excimer laser delivery system for astigmatic and hyperopic photorefractive surgery

NASA Astrophysics Data System (ADS)

Beck, Rasmus; Foerster, Werner

1994-06-01

Ablation of corneal tissue with excimer laser light is an effective way to correct refractive errors of the eye. For this purpose a beam-stop (iris diaphragm or interchangeable masks) is illuminated by the laser radiation. The beam-stop is imaged onto the cornea, and circular or elliptic ablations are produced. The computer-controlled process varies the diameter of the ablation area in a way that the inner portions of the treatment zone receive more laser energy than the outer portions, thus flattening the curvature of the refractive surface. For the treatment of hyperopia, the outer portions of the ablation area receive more laser energy to steepen the surface profile of the cornea. The beam delivery system employs several sets of circular, elliptic and ring shaped masks which are etched into a stainless-steel tape.

Self-organized centripetal movement of corneal epithelium in the absence of external cues

NASA Astrophysics Data System (ADS)

Lobo, Erwin P.; Delic, Naomi C.; Richardson, Alex; Raviraj, Vanisri; Halliday, Gary M.; di Girolamo, Nick; Myerscough, Mary R.; Lyons, J. Guy

2016-08-01

Maintaining the structure of the cornea is essential for high-quality vision. In adult mammals, corneal epithelial cells emanate from stem cells in the limbus, driven by an unknown mechanism towards the centre of the cornea as cohesive clonal groups. Here we use complementary mathematical and biological models to show that corneal epithelial cells can self-organize into a cohesive, centripetal growth pattern in the absence of external physiological cues. Three conditions are required: a circumferential location of stem cells, a limited number of cell divisions and mobility in response to population pressure. We have used these complementary models to provide explanations for the increased rate of centripetal migration caused by wounding and the potential for stem cell leakage to account for stable transplants derived from central corneal tissue, despite the predominantly limbal location of stem cells.

The Impact of Inoculation Parameters on the Pathogenesis of Contact Lens–Related Infectious Keratitis

PubMed Central

Tam, Connie; Mun, James J.; Evans, David J.

2010-01-01

Purpose. Contact lens wear predisposes to Pseudomonas aeruginosa keratitis, but the mechanisms involved remain unclear. An in vivo model was used to study lens inoculation conditions enabling disease. Methods. Custom-made hydrogel contact lenses were fitted to rats after incubation in P. aeruginosa approximately 1011 cfu/mL (3 hours) or approximately 103 cfu/mL (24 hours). Another group was inadvertently inoculated with a suction pen previously used with high inocula, but rinsed in ethanol and stored dry (6 months). Some corneas were tissue paper–blotted to cause fluorescein staining before lens fitting. Contralateral eyes were untreated. Twenty-four hours after disease detection, lenses were transferred to naive rats or examined by confocal microscopy before homogenization to quantify viable bacteria. After lens removal, corneas were washed to collect nonadherent bacteria and were analyzed by immunohistochemistry. Results. All eyes challenged with unworn contaminated lenses developed keratitis after approximately 7 to 10 days. Disease delay and severity were unaffected by inoculum parameters or tissue blotting but occurred sooner with lenses transferred from infected eyes (∼2 days). Worn lenses and corneal washes contained infecting bacteria. Posterior, not anterior, lens surfaces harbored P. aeruginosa biofilms that penetrated the lens matrix. Diseased corneas showed an infiltration of phagocytes and T-lymphocytes. Conclusions. P. aeruginosa induces keratitis in this lens-wearing model after a single inoculation. Delayed disease onset was interesting considering the greater keratitis risk during extended wear. Infection did not require the disruption of corneal barrier function before lens wear and occurred without exposure to lens care solutions. The data suggest that keratitis involves biofilm formation or other bacterial adaptations in vivo. PMID:20130275

Nitrogen Mustard-Induced Corneal Injury Involves DNA Damage and Pathways Related to Inflammation, Epithelial-Stromal Separation, and Neovascularization.

PubMed

Goswami, Dinesh G; Tewari-Singh, Neera; Dhar, Deepanshi; Kumar, Dileep; Agarwal, Chapla; Ammar, David A; Kant, Rama; Enzenauer, Robert W; Petrash, J Mark; Agarwal, Rajesh

2016-02-01

To evaluate the toxic effects and associated mechanisms in corneal tissue exposed to the vesicating agent, nitrogen mustard (NM), a bifunctional alkylating analog of the chemical warfare agent sulfur mustard. Toxic effects and associated mechanisms were examined in maximally affected corneal tissue using corneal cultures and human corneal epithelial (HCE) cells exposed to NM. Analysis of ex vivo rabbit corneas showed that NM exposure increased apoptotic cell death, epithelial thickness, epithelial-stromal separation, and levels of vascular endothelial growth factor, cyclooxygenase 2, and matrix metalloproteinase-9. In HCE cells, NM exposure resulted in a dose-dependent decrease in cell viability and proliferation, which was associated with DNA damage in terms of an increase in p53 ser15, total p53, and H2A.X ser139 levels. NM exposure also induced caspase-3 and poly ADP ribose polymerase cleavage, suggesting their involvement in NM-induced apoptotic death in the rabbit cornea and HCE cells. Similar to rabbit cornea, NM exposure caused an increase in cyclooxygenase 2, matrix metalloproteinase-9, and vascular endothelial growth factor levels in HCE cells, indicating a role of these molecules and related pathways in NM-induced corneal inflammation, epithelial-stromal separation, and neovascularization. NM exposure also induced activation of activator protein 1 transcription factor proteins and upstream signaling pathways including mitogen-activated protein kinases and Akt protein kinase, suggesting that these could be key factors involved in NM-induced corneal injury. Results from this study provide insight into the molecular targets and pathways that could be involved in NM-induced corneal injuries laying the background for further investigation of these pathways in vesicant-induced ocular injuries, which could be helpful in the development of targeted therapies.

Nitrogen mustard-induced corneal injury involves DNA damage and pathways related to inflammation, epithelial-stromal separation and neovascularization

PubMed Central

Goswami, Dinesh G; Tewari-Singh, Neera; Dhar, Deepanshi; Kumar, Dileep; Agarwal, Chapla; Ammar, David A; Kant, Rama; Enzenauer, Robert W; Petrash, J Mark; Agarwal, Rajesh

2015-01-01

Purpose To evaluate the toxic effects and associated mechanisms in corneal tissue exposed to vesicating agent, nitrogen mustard (NM), a bi-functional alkylating analog of chemical warfare agent sulfur mustard (SM). Methods Toxic effects and associated mechanisms were examined in maximal affected corneal tissue employing corneal cultures and human corneal epithelial (HCE) cells exposed to nitrogen mustard (NM). Results Analysis of ex vivo rabbit corneas showed that NM exposure increased apoptotic cell death, epithelial thickness, epithelial-stromal separation and levels of VEGF, COX-2 and MMP-9. In HCE cells, NM exposure resulted in a dose-dependent decrease in cell viability and proliferation, which was associated with DNA damage in terms of an increase in p53 ser15, total p53 and H2A.X ser139 levels. NM exposure also induced caspase-3 and PARP cleavage, suggesting their involvement in NM-induced apoptotic death in rabbit cornea and HCE cells. Similar to rabbit cornea, NM exposure caused an increase in COX-2, MMP-9 and VEGF levels in HCE cells, indicating a role of these molecules and related pathways in NM-induced corneal inflammation, epithelial-stromal separation and neovascularization. NM exposure also induced activation of AP-1 transcription factor proteins and upstream signaling pathways including MAPKs and Akt, suggesting that these could be key factors involved in NM-induced corneal injury. Conclusion Results from this study provide insight into the molecular targets and pathways that could be involved in NM-induced corneal injuries laying the background for further investigation of these pathways in vesicant–induced ocular injuries, which could be helpful in the development of targeted therapies. PMID:26555588

Photokeratitis induced by ultraviolet radiation in travelers: A major health problem

PubMed Central

Izadi, M; Jonaidi-Jafari, N; Pourazizi, M; Alemzadeh-Ansari, MH; Hoseinpourfard, MJ

2018-01-01

Ultraviolet (UV) irradiation is one of the several environmental hazards that may cause inflammatory reactions in ocular tissues, especially the cornea. One of the important factors that affect how much ultraviolet radiation (UVR) humans are exposed to is travel. Hence, traveling is considered to include a more acute UVR effect, and ophthalmologists frequently evaluate and manage the ocular manifestations of UV irradiation, including UV-induced keratitis. The purpose of this paper is to provide an evidence-based analysis of the clinical effect of UVR in ocular tissues. An extensive review of English literature was performed to gather all available articles from the National Library of Medicine PubMed database of the National Institute of Health, the Ovid MEDLINE database, Scopus, and ScienceDirect that had studied the effect of UVR on the eye and its complications, between January 1970 and June 2014. The results show that UVR at 300 nm causes apoptosis in all three layers of the cornea and induces keratitis. Apoptosis in all layers of the cornea occurs 5 h after exposure. The effect of UVR intensity on the eye can be linked to numerous factors, including solar elevation, time of day, season, hemisphere, clouds and haze, atmospheric scattering, atmospheric ozone, latitude, altitude, longitudinal changes, climate, ground reflection, and geographic directions. The most important factor affecting UVR reaching the earth's surface is solar elevation. Currently, people do not have great concern over eye protection. The methods of protection against UVR include avoiding direct sunlight exposure, using UVR-blocking eyewear (sunglasses or contact lenses), and wearing hats. Hence, by identifying UVR intensity factors, eye protection factors, and public education, especially in travelers, methods for safe traveling can be identified. PMID:29067921

Photokeratitis induced by ultraviolet radiation in travelers: A major health problem.

PubMed

Izadi, M; Jonaidi-Jafari, N; Pourazizi, M; Alemzadeh-Ansari, M H; Hoseinpourfard, M J

2018-01-01

Ultraviolet (UV) irradiation is one of the several environmental hazards that may cause inflammatory reactions in ocular tissues, especially the cornea. One of the important factors that affect how much ultraviolet radiation (UVR) humans are exposed to is travel. Hence, traveling is considered to include a more acute UVR effect, and ophthalmologists frequently evaluate and manage the ocular manifestations of UV irradiation, including UV-induced keratitis. The purpose of this paper is to provide an evidence-based analysis of the clinical effect of UVR in ocular tissues. An extensive review of English literature was performed to gather all available articles from the National Library of Medicine PubMed database of the National Institute of Health, the Ovid MEDLINE database, Scopus, and ScienceDirect that had studied the effect of UVR on the eye and its complications, between January 1970 and June 2014. The results show that UVR at 300 nm causes apoptosis in all three layers of the cornea and induces keratitis. Apoptosis in all layers of the cornea occurs 5 h after exposure. The effect of UVR intensity on the eye can be linked to numerous factors, including solar elevation, time of day, season, hemisphere, clouds and haze, atmospheric scattering, atmospheric ozone, latitude, altitude, longitudinal changes, climate, ground reflection, and geographic directions. The most important factor affecting UVR reaching the earth's surface is solar elevation. Currently, people do not have great concern over eye protection. The methods of protection against UVR include avoiding direct sunlight exposure, using UVR-blocking eyewear (sunglasses or contact lenses), and wearing hats. Hence, by identifying UVR intensity factors, eye protection factors, and public education, especially in travelers, methods for safe traveling can be identified.

Corneal Resistance to Keratolysis After Collagen Crosslinking With Rose Bengal and Green Light.

PubMed

Fadlallah, Ali; Zhu, Hong; Arafat, Samer; Kochevar, Irene; Melki, Samir; Ciolino, Joseph B

2016-12-01

The purpose of this study was to evaluate the resistance to degradation by collagenase A of corneas that have been crosslinked with Rose Bengal and green light (RGX). The ex vivo crosslinking procedure was performed on enucleated rabbit corneas. Corneas were deepithelialized after applying 30% alcohol. Corneas were stained with Rose Bengal (RB, 0.1%) for 2 minutes and then exposed to green light (532 nm) at 0.25 W/cm2 for times to deliver doses of 50, 100, 150, or 200 J/cm2 (n = 5 per group). Five corneas were pretreated with riboflavin solution (0.1% riboflavin) for 15 minutes and irradiated with ultraviolet A (UVA) light (370 nm, 3 mW/cm2) for 30 minutes. Five corneas underwent only de-epithelialization and were otherwise untreated. Five corneas were stained with RB without light exposure. The central corneas of each group was removed with a 8.5-mm trephine and incubated at 37°C in 0.3% collagenase A solution. Time to dissolution of each cornea was compared across treatments. Corneas treated with RGX were treated with light fluences of 50, 100, 150, and 200 J/cm2; these corneas dissolved completely at 8.3 ± 1.2, 11.1 ± 1.4, 12.4 ± 1.7, and 15.7 ± 1.8 hours, respectively. Corneas treated by riboflavin and UVA light dissolved at 15.7 ± 1.7 hours, and nontreated corneas dissolved at 6.1 ± 1.3 hours. Corneas treated with only RB (no green light) dissolved at 9.3 ± 1.7 hours. Compared with the untreated corneas, all of the RB groups and the riboflavin-UVA-treated group of corneas degraded statistically significantly slower than untreated corneas (P < 0.05). Crosslinking with RGX increased corneal resistance to digestion by collagenase comparable to that produced by riboflavin and UVA treatment.

Micro-processing of polymers and biological materials using high repetition rate femtosecond laser pulses

NASA Astrophysics Data System (ADS)

Ding, Li

High repetition rate femtosecond laser micro-processing has been applied to ophthalmological hydrogel polymers and ocular tissues to create novel refractive and diffractive structures. Through the optimization of laser irradiation conditions and material properties, this technology has become feasible for future industrial applications and clinical practices. A femtosecond laser micro-processing workstation has been designed and developed. Different experimental parameters of the workstation such as laser pulse duration, focusing lens, and translational stages have been described and discussed. Diffractive gratings and three-dimensional waveguides have been fabricated and characterized in hydrogel polymers, and refractive index modifications as large as + 0.06 have been observed within the laser-irradiated region. Raman spectroscopic studies have shown that our femtosecond laser micro-processing induces significant thermal accumulation, resulting in a densification of the polymer network and increasing the localized refractive index of polymers within the laser irradiated region. Different kinds of dye chromophores have been doped in hydrogel polymers to enhance the two-photon absorption during femtosecond laser micro-processing. As the result, laser scanning speed can be greatly increased while the large refractive index modifications remain. Femtosecond laser wavelength and pulse energy as well as water and dye concentration of the hydrogels are optimized. Lightly fixed ocular tissues such as corneas and lenses have been micro-processed by focused femtosecond laser pulses, and refractive index modifications without any tissue-breakdown are observed within the stromal layer of the corneas and the cortex of the lenses. Living corneas are doped with Sodium Fluorescein to increase the two-photon absorption during the laser micro-processing, and laser scanning speed can be greatly increased while inducing large refractive index modifications. No evidence of cell death has been observed in or around the laser-induced refractive index modification regions. These results support the notion that femtosecond laser micro-processing method may be an excellent means of altering the refraction or higher order aberration content of corneal tissue without cell death and short-term tissue damage, and has been named as Intra-tissue Refractive Index Shaping (IRIS). The femtosecond laser micro-processing workstation has also been employed for laser transfection of single defined cells. Some preliminary results suggest that this method can be used to trace individual cells and record their biological and morphological evolution, which is quite promising in many biomedical applications especially in immunology science. In conclusion, high repetition rate femtosecond laser micro-processing has been employed to fabricate microstructures in ophthalmological hydrogels and ocular tissues. Its unique three-dimensional capability over transparent materials and biological media makes it a powerful tool and will greatly impact the future of laser material-processing.

A portable device for measuring donor corneal transparency in eye banks.

PubMed

Parekh, Mohit; Ferrari, Stefano; Ruzza, Alessandro; Pugliese, Mariarosaria; Ponzin, Diego; Salvalaio, Gianni

2014-03-01

To develop a portable device for measuring the donor corneal transparency and validate its efficacy for corneal evaluation in the eye-banks and for research. The transparency device (TD) has a light source, a detachable system for corneal insertion and a base for light transmission. The probe detects the transmitted light which is measured by a lux-meter. A contact lens was set as 'control' to reduce the light scattering concern, an empty petri-plate as 'blank' and the cornea as 'sample'. Two experts and non-experts (masked) observed the corneas for subjective analysis which was then compared using the TD. The parameters observed were scars, foreign-body, stromal-deformities, folds, thickness and opacity which were then converted to a relative overall percentage by the observer. Twenty corneas were evaluated for correlation, five tissues to obtain standard-deviation and twenty-four pairs for a comparative study. Experts mimicked the eye-banks with long-term experience while non-experts mimicked the emerging eye-banks. Subjective values by the experts closely resembled the measurements by TD. The average correlation between the experts and the non-experts to TD was 0.985 and 0.960 respectively. TD showed higher reproducibility than experts followed by the non-experts. The comparative study showed that increase in thickness reduces the transparency. TD is portable, easy, efficient, maintains sterility and less expensive hence the emerging eye-banks and researchers can use to raise their standards and evaluate the transparency for in vitro tests and comparative studies. The suitable transparency for the cornea deemed for clinical applications was found to be >75 %.

Corneal tissue water content mapping with THz imaging: preliminary clinical results (Conference Presentation)

NASA Astrophysics Data System (ADS)

Sung, Shijun; Bajwa, Neha; Deng, Sophie X.; Taylor, Zachary; Grundfest, Warren

2016-03-01

Well-regulated corneal water content is critical for ocular health and function and can be adversely affected by a number of diseases and injuries. Current clinical practice limits detection of unhealthy corneal water content levels to central corneal thickness measurements performed by ultrasound or optical coherence tomography. Trends revealing increasing or decreasing corneal thickness are fair indicators of corneal water content by individual measurements are highly inaccurate due to the poorly understood relationship between corneal thickness and natural physiologic variation. Recently the utility of THz imaging to accuarately measure corneal water content has been explored on with rabbit models. Preliminary experiments revealed that contact with dielectric windows confounded imaging data and made it nearly impossible to deconvolve thickness variations due to contact from thickness variations due to water content variation. A follow up study with a new optical design allowed the acquisition of rabbit data and the results suggest that the observed, time varying contrast was due entirely to the water dynamics of the cornea. This paper presents the first ever in vivo images of human cornea. Five volunteers with healthy cornea were recruited and their eyes were imaged three times over the course of a few minutes with our novel imaging system. Noticeable changes in corneal reflectivity were observed and attributed to the drying of the tear film. The results suggest that clinically compatible, non-contact corneal imaging is feasible and indicate that signal acquired from non-contact imaging of the cornea is a complicated coupling of stromal water content and tear film.

Noninvasive measurement of wave speed of porcine cornea in ex vivo porcine eyes for various intraocular pressures.

PubMed

Zhou, Boran; Sit, Arthur J; Zhang, Xiaoming

2017-11-01

The objective of this study was to extend an ultrasound surface wave elastography (USWE) technique for noninvasive measurement of ocular tissue elastic properties. In particular, we aim to establish the relationship between the wave speed of cornea and the intraocular pressure (IOP). Normal ranges of IOP are between 12 and 22mmHg. Ex vivo porcine eye balls were used in this research. The porcine eye ball was supported by the gelatin phantom in a testing container. Some water was pour into the container for the ultrasound measurement. A local harmonic vibration was generated on the side of the eye ball. An ultrasound probe was used to measure the wave propagation in the cornea noninvasively. A 25 gauge butterfly needle was inserted into the vitreous humor of the eye ball under the ultrasound imaging guidance. The needle was connected to a syringe. The IOP was obtained by the water height difference between the water level in the syringe and the water level in the testing container. The IOP was adjusted between 5mmHg and 30mmHg with a 5mmHg interval. The wave speed was measured at each IOP for three frequencies of 100, 150 and 200Hz. Finite element method (FEM) was used to simulate the wave propagation in the corneal according to our experimental setup. A linear viscoelastic FEM model was used to compare the experimental data. Both the experiments and the FEM analyses showed that the wave speed of cornea increased with IOP. Copyright © 2017 Elsevier B.V. All rights reserved.

Confocal microscopy and electrophysiological study of single patient corneal endothelium cell cultures

NASA Astrophysics Data System (ADS)

Tatini, Francesca; Rossi, Francesca; Coppi, Elisabetta; Magni, Giada; Fusco, Irene; Menabuoni, Luca; Pedata, Felicita; Pugliese, Anna Maria; Pini, Roberto

2016-04-01

The characterization of the ion channels in corneal endothelial cells and the elucidation of their involvement in corneal pathologies would lead to the identification of new molecular target for pharmacological treatments and to the clarification of corneal physiology. The corneal endothelium is an amitotic cell monolayer with a major role in preserving corneal transparency and in regulating the water and solute flux across the posterior surface of the cornea. Although endothelial cells are non-excitable, they express a range of ion channels, such as voltage-dependent Na+ channels and K+ channels, L-type Ca2 channels and many others. Interestingly, purinergic receptors have been linked to a variety of conditions within the eye but their presence in the endothelium and their role in its pathophysiology is still uncertain. In this study, we were able to extract endothelial cells from single human corneas, thus obtaining primary cultures that represent the peculiarity of each donor. Corneas were from tissues not suitable for transplant in patients. We characterized the endothelial cells by confocal microscopy, both within the intact cornea and in the primary endothelial cells cultures. We also studied the functional role of the purinergic system (adenosine, ATP and their receptors) by means of electrophysiological recordings. The experiments were performed by patch clamp recordings and confocal time-lapse microscopy and our results indicate that the application of purinergic compounds modulates the amplitude of outward currents in the isolated endothelial cells. These findings may lead to the proposal of new therapies for endothelium-related corneal diseases.

Alkaptonuria: A case report.

PubMed

Damarla, Nirupama; Linga, Prathima; Goyal, Mallika; Tadisina, Sanjay Reddy; Reddy, G Satyanarayana; Bommisetti, Hymavathi

2017-06-01

Alkaptonuria is a rare inborn error of metabolism with autosomal recessive inheritance with a mutation in homogentisate 1,2-dioxygenase. It results in accumulation of homogentisic acid in connective tissues (ochronosis). Most common ocular manifestations are bluish-black discoloration of the conjunctiva, cornea, and sclera. In this case report, a 39-year-old Indian male patient with additional ocular features in the retina is described.

Alkaptonuria: A case report

PubMed Central

Damarla, Nirupama; Linga, Prathima; Goyal, Mallika; Tadisina, Sanjay Reddy; Reddy, G Satyanarayana; Bommisetti, Hymavathi

2017-01-01

Alkaptonuria is a rare inborn error of metabolism with autosomal recessive inheritance with a mutation in homogentisate 1,2-dioxygenase. It results in accumulation of homogentisic acid in connective tissues (ochronosis). Most common ocular manifestations are bluish-black discoloration of the conjunctiva, cornea, and sclera. In this case report, a 39-year-old Indian male patient with additional ocular features in the retina is described. PMID:28643719

Contact diode laser application through a fiberoptic cutting tip.

PubMed

Peyman, G A; Cruz, S A; Ruiz-Lapuente, C

1991-01-01

We applied diode laser energy through a fiberoptic probe to cut ocular structures. The diameter of the probe's tip was 100 microns. The amount of energy used for cutting the lid, the cornea, or for perforation of the glove was 2.5 watts in the continuous mode. The incised border demonstrated coagulation into the healthy tissue ranging from 10 to 50 microns.

Integrated instrument for dynamic light scattering and natural fluorescence measurements

NASA Astrophysics Data System (ADS)

Rovati, Luigi; Pollonini, Luca; Ansari, Rafat R.

2001-06-01

Over the past two decades, great efforts have been made in ophthalmology to use optical techniques based on dynamic light scattering and tissue natural fluorescence for early (at molecular level) diagnosis of ocular pathologies. In our previous studies, the relationship between the corneal AF and DLS decay widths of ocular tissues were established by performing measurements on diabetes mellitus patients. In those studies, corneal AF mean intensities were significantly correlated with DLS decay width measurements for each diabetic retinopathy grade in the vitreous and in the cornea. This suggested that the quality of the diagnosis could be significantly improved by properly combining these two powerful techniques into a single instrument. Our approach is based on modifying a commercial scanning ocular fluorometer (Fluorotron Master, Ocumetrics Inc., CA, USA) to include both techniques in the same scanning unit. This configuration provides both DLS and AF real time measurements from the same ocular volume: they can be located in each section of the optical axis of the eye from the cornea to the retina. In this paper, the optical setup of the new system is described and preliminary in-vitro and in-vivo measurements are presented.

The Pathogenesis of Staphylococcus aureus Eye Infections

PubMed Central

2018-01-01

Staphylococcus aureus is a major pathogen of the eye able to infect the tear duct, eyelid, conjunctiva, cornea, anterior and posterior chambers, and the vitreous chamber. Of these infections, those involving the cornea (keratitis) or the inner chambers of the eye (endophthalmitis) are the most threatening because of their potential to cause a loss in visual acuity or even blindness. Each of these ocular sites is protected by the constitutive expression of a variety of antimicrobial factors and these defenses are augmented by a protective host response to the organism. Such infections often involve a predisposing factor that weakens the defenses, such as the use of contact lenses prior to the development of bacterial keratitis or, for endophthalmitis, the trauma caused by cataract surgery or intravitreal injection. The structural carbohydrates of the bacterial surface induce an inflammatory response able to reduce the bacterial load, but contribute to the tissue damage. A variety of bacterial secreted proteins including alpha-toxin, beta-toxin, gamma-toxin, Panton-Valentine leukocidin and other two-component leukocidins mediate tissue damage and contribute to the induction of the inflammatory response. Quantitative animal models of keratitis and endophthalmitis have provided insights into the S. aureus virulence and host factors active in limiting such infections. PMID:29320451

Eye-bank Preparation of Endothelial Tissue

PubMed Central

Boynton, Grace E.; Woodward, Maria A.

2014-01-01

Purpose of review Eyebank preparation of endothelial tissue for keratoplasty continues to evolve. While eye bank personnel have become comfortable and competent at Descemet Stripping Automated Endothelial Keratoplasty (DSAEK) tissue preparation and tissue transport, optimization of preparation methods continues. Surgeons and eye bank personnel should be up to date on the research in the field. As surgeons transition to Descemet Membrane Endothelial Keratoplasty (DMEK), eye banks have risen to the challenge of preparing tissue. Eye banks are refining their DMEK preparation and transport techniques Recent findings This article covers refinements to DSAEK tissue preparation, innovations to prepare DMEK tissue, and nuances to improve donor cornea tissue quality. Summary As eye bank supplied corneal tissue is the main source of tissue for many corneal surgeons, it is critical to stay informed about tissue handling and preparation. Ultimately the surgeon is responsible for the transplantation, so involvement of clinicians in eye banking practices and advocacy for pursuing meaningful research in this area will benefit clinical patient outcomes. PMID:24837574

Reduction in Histone H3 Acetylation and Chromatin Remodeling in Corneas of Alloxan-Induced Diabetic Rats.

PubMed

Herencia-Bueno, Karina E; Aldrovani, Marcela; Crivelaro, Roberta M; Thiesen, Roberto; Barros-Sobrinho, Alexandre A F; Claros-Chacaltana, Flor D Y; Padua, Ivan R M; Santos, Daniela M; Laus, José L

2018-05-01

To evaluate acetylation of histone H3, chromatin remodeling, nuclear size and shape, DNA ploidy, and distribution of nucleolus organizing regions (NORs) in corneal epithelial and stromal cells of diabetic and nondiabetic rats. Diabetes was induced by a single intraperitoneal injection of alloxan. All diabetic rats (n = 20) included in the study had 4 weeks of moderate-to-severe hyperglycemia (plasma glucose levels >400 mg/dL). Acetylated histone H3 levels were quantified in corneal tissue using a colorimetric assay. Chromatin remodeling, nuclear sizes (area/perimeter) and shapes (circularity), and DNA ploidies were evaluated from Feulgen-stained tissue sections using video image analysis. Distributions of NORs were studied in tissue sections impregnated with silver ions. Ophthalmic clinical parameters, including corneal sensitivity, were investigated. Twenty nondiabetic rats were used as controls. Acetylation of histone H3 was reduced in the corneas of the diabetic rats. Nuclei in corneal epithelial cells of diabetic rats compacted chromatin, increased in size, modified their shapes, and elevated DNA ploidy. The only nuclear change observed in the corneal stromal cells of diabetic rats was chromatin decompaction. The size of the silver-stained NOR did not differ between the study samples. The corneal sensitivity in diabetic rats was 51.8% lower than that in nondiabetic rats. The results of this study show that alloxan-induced diabetes altered the histone H3 acetylation pattern and compromised the chromatin supraorganization in corneal tissue/cells. Continued research is needed to understand the clinical and morphofunctional significance of changes in corneal cell nuclei of diabetic individuals.

Functional fabrication of recombinant human collagen-phosphorylcholine hydrogels for regenerative medicine applications.

PubMed

Mirazul Islam, M; Cėpla, Vytautas; He, Chaoliang; Edin, Joel; Rakickas, Tomas; Kobuch, Karin; Ruželė, Živilė; Bruce Jackson, W; Rafat, Mehrdad; Lohmann, Chris P; Valiokas, Ramūnas; Griffith, May

2015-01-01

The implant-host interface is a critical element in guiding tissue or organ regeneration. We previously developed hydrogels comprising interpenetrating networks of recombinant human collagen type III and 2-methacryloyloxyethyl phosphorylcholine (RHCIII-MPC) as substitutes for the corneal extracellular matrix that promote endogenous regeneration of corneal tissue. To render them functional for clinical application, we have now optimized their composition and thereby enhanced their mechanical properties. We have demonstrated that such optimized RHCIII-MPC hydrogels are suitable for precision femtosecond laser cutting to produce complementing implants and host surgical beds for subsequent tissue welding. This avoids the tissue damage and inflammation associated with manual surgical techniques, thereby leading to more efficient healing. Although we previously demonstrated in clinical testing that RHCIII-based implants stimulated cornea regeneration in patients, the rate of epithelial cell coverage of the implants needs improvement, e.g. modification of the implant surface. We now show that our 500μm thick RHCIII-MPC constructs comprising over 85% water are suitable for microcontact printing with fibronectin. The resulting fibronectin micropatterns promote cell adhesion, unlike the bare RHCIII-MPC hydrogel. Interestingly, a pattern of 30μm wide fibronectin stripes enhanced cell attachment and showed the highest mitotic rates, an effect that potentially can be utilized for faster integration of the implant. We have therefore shown that laboratory-produced mimics of naturally occurring collagen and phospholipids can be fabricated into robust hydrogels that can be laser profiled and patterned to enhance their potential function as artificial substitutes of donor human corneas. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Differential distribution of blood and lymphatic vessels in the murine cornea.

PubMed

Ecoiffier, Tatiana; Yuen, Don; Chen, Lu

2010-05-01

Because of its unique characteristics, the cornea has been widely used for blood and lymphatic vessel research. However, whether limbal or corneal vessels are evenly distributed under normal or inflamed conditions has never been studied. The purpose of this study was to investigate this question and to examine whether and how the distribution patterns change during corneal inflammatory lymphangiogenesis (LG) and hemangiogenesis (HG). Corneal inflammatory LG and HG were induced in two most commonly used mouse strains, BALB/c and C57BL/6 (6-8 weeks of age), by a standardized two-suture placement model. Oriented flat-mount corneas together with the limbal tissues were used for immunofluorescence microscope studies. Blood and lymphatic vessels under normal and inflamed conditions were analyzed and quantified to compare their distributions. The data demonstrate, for the first time, greater distribution of both blood and lymphatic vessels in the nasal side in normal murine limbal areas. This nasal-dominant pattern was maintained during corneal inflammatory LG, whereas it was lost for HG. Blood and lymphatic vessels are not evenly distributed in normal limbal areas. Furthermore, corneal LG and HG respond differently to inflammatory stimuli. These new findings will shed some light on corneal physiology and pathogenesis and on the development of experimental models and therapeutic strategies for corneal diseases.

VEGF-A expression by HSV-1–infected cells drives corneal lymphangiogenesis

PubMed Central

Wuest, Todd R.

2010-01-01

Inflammatory lymphangiogenesis plays a crucial role in the development of inflammation and transplant rejection. The mechanisms of inflammatory lymphangiogenesis during bacterial infection, toll-like receptor ligand administration, and wound healing are well characterized and depend on ligands for the vascular endothelial grow factor receptor (VEGFR) 3 that are produced by infiltrating macrophages. But inflammatory lymphangiogenesis in nonlymphoid tissues during chronic viral infection is unstudied. Herpes simplex virus 1 (HSV-1) infection of the cornea is a leading cause of blindness and depends on aberrant host immune responses to antigen within the normally immunologically privileged cornea. We report that corneal HSV-1 infection drives lymphangiogenesis and that corneal lymphatics persist past the resolution of infection. The mechanism of HSV-1–induced lymphangiogenesis was distinct from the described mechanisms of inflammatory lymphangiogenesis. HSV-1–elicited lymphangiogenesis was strictly dependent on VEGF-A/VEGFR-2 signaling but not on VEGFR-3 ligands. Macrophages played no role in the induction of lymphangiogenesis and were not a detectable source of VEGF-A. Rather, using VEGF-A reporter transgenic mice, we have identified infected epithelial cells as the primary source of VEGF-A during HSV-1 infection. Our results indicate that HSV-1 directly induces vascularization of the cornea through up-regulation of VEGF-A expression. PMID:20026662

Measuring the Refractive Index of Bovine Corneal Stromal Cells Using Quantitative Phase Imaging

PubMed Central

Gardner, Steven J.; White, Nick; Albon, Julie; Knupp, Carlo; Kamma-Lorger, Christina S.; Meek, Keith M.

2015-01-01

The cornea is the primary refractive lens in the eye and transmits >90% of incident visible light. It has been suggested that the development of postoperative corneal haze could be due to an increase in light scattering from activated corneal stromal cells. Quiescent keratocytes are thought to produce crystallins that match the refractive index of their cytoplasm to the surrounding extracellular material, reducing the amount of light scattering. To test this, we measured the refractive index (RI) of bovine corneal stromal cells, using quantitative phase imaging of live cells in vitro, together with confocal microscopy. The RI of quiescent keratocytes (RI = 1.381 ± 0.004) matched the surrounding matrix, thus supporting the hypothesis that keratocyte cytoplasm does not scatter light in the normal cornea. We also observed that the RI drops after keratocyte activation (RI = 1.365 ± 0.003), leading to a mismatch with the surrounding intercellular matrix. Theoretical scattering models showed that this mismatch would reduce light transmission in the cornea. We conclude that corneal transparency depends on the matching of refractive indices between quiescent keratocytes and the surrounding tissue, and that after surgery or wounding, the resulting RI mismatch between the activated cells and their surrounds significantly contributes to light scattering. PMID:26488650

Improved Ocular Delivery of Nepafenac by Cyclodextrin Complexation.

PubMed

Shelley, Haley; Grant, Makenzie; Smith, Forrest T; Abarca, Eva M; Jayachandra Babu, R

2018-06-13

Nepafenac is a nonsteroidal anti-inflammatory drug (NSAID), currently only available as 0.1% ophthalmic suspension (Nevanac®). This study utilized hydroxypropyl-β-cyclodextrin (HPBCD) to increase the water solubility and trans-corneal permeation of nepafenac. The nepafenac-HPBCD complexation in the liquid and solid states were confirmed by phase solubility, differential scanning calorimetry (DSC), X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), and nuclear magnetic resonance spectroscopy (NMR) analyses. Nepafenac 0.1% ophthalmic solution was formulated using HPBCD (same pH and osmolality as that of Nevanac®) and pig eye trans-corneal permeation was studied versus Nevanac®. Furthermore, nepafenac content in cornea, sclera, iris, lens, aqueous humor, choroid, ciliary body, retina, and vitreous humor was studied in a continuous isolated pig eye perfusion model in comparison to the suspension and Nevanac®. Permeation studies using porcine corneas revealed that the solution formulation had a permeation rate 18 times higher than Nevanac®. Furthermore, the solution had 11 times higher corneal retention than Nevanac®. Drug distribution studies using porcine eyes revealed that the solution formulation enables detectable levels in various ocular tissues while the drug was undetectable by Nevanac®. The ocular solution formulation had a significantly higher drug concentration in the cornea compared to the suspension or Nevanac®.

Mast Cell Activation Protects Cornea by Promoting Neutrophil Infiltration via Stimulating ICAM-1 and Vascular Dilation in Fungal Keratitis.

PubMed

Xie, Yanting; Zhang, Hongmin; Liu, Susu; Chen, Guoming; He, Siyu; Li, Zhijie; Wang, Liya

2018-05-30

The role of mast cells (MCs) in fungal infection is largely unknown. This study was to explore a protective role and mechanism of MCs in fungal keratitis. Experimental fungal keratitis (FK) mouse model was developed. Mice untreated (UT) or receiving corneal wound without fungal infection (Mock) were used as controls. Large number of connective tissue MCs was found in normal mice. MC activation with degranulation was largely observed, and the percentage of degranulated/total cells was high in FK. Dilated limbal vasculature with increased permeability, as well as largely infiltrated neutrophils with stimulated ICAM-1 protein levels were observed in corneas of FK mice, when compared with Mock and UT mice. Interestingly, pretreatment with cromolyn sodium (Block) significantly blocked MC degranulation, dramatically suppressed vascular dilation and permeability, and markedly reduced neutrophil infiltration with lower ICAM-1 levels in FK mice at 6-24 hours. Furthermore, the Block mice manifested prolonged disease course, increased pathological damage, and vigorous fungus growth, with much higher corneal perforation rate than FK mice at 72 h. These findings reveal a novel phenomenon that MCs play a vital role in protecting cornea against fungal infection through degranulation that promotes neutrophil infiltration via stimulating ICAM-1 production and limbal vascular dilation and permeability.

The management of corneal trauma: advances in the past twenty-five years.

PubMed

Macsai, M S

2000-09-01

Over the past quarter century, advances in our understanding of corneal anatomy, physiology, and wound healing have all played an integral role in the management of corneal trauma. As the etiologies of corneal trauma have changed, so has our understanding of the impact of injury on corneal function as it relates to visual rehabilitation. Numerous new classes of antibiotics, antiinflammatory agents, and tissue adhesives have emerged. Occlusive therapy has advanced from simple pressure patching bandage soft contact lenses and collagen shields. Surgical instrumentation, operating microscopes, viscoelastic substances, and suture materials have all improved the outcomes of corneal trauma repair. Improved understanding of the refractive properties of the cornea through topography and alternative suture techniques has helped us restore the natural corneal curvature and visual outcomes. Consequently, in the last quarter of this century our therapeutic approaches to cornea trauma, both medical and surgical, have improved.

Keratoprosthesis: preliminary results of an artificial corneal button as a full-thickness implant in the rabbit model.

PubMed

Hicks, C R; Chirila, T V; Dalton, P D; Clayton, A B; Vijayasekaran, S; Crawford, G J; Constable, I J

1996-08-01

To develop a prototype artificial cornea and evaluate it in the rabbit model. Hydrogel core-and-skirt keratoprostheses were made and were inserted as full-thickness implants covered with conjunctival flaps in the right eyes of eight rabbits. Peroperative complications related to inadequate mechanical strength led to failure in the early postoperative period in three animals, one was euthanased for an unrelated reason and the remaining four have been successful for up to 16 weeks' follow-up. Full-thickness implantation of an artificial cornea, analogous to penetrating keratoplasty, has been achieved in the rabbit model. Histological findings confirm that integration of the prosthesis with host tissue occurs. The main complications encountered in this preliminary series were related to inadequate strength of the sponge skirt of this prototype device. Work in our laboratories is now concentrated upon improving the mechanical qualities of the hydrogel skirt and on the enhancement of biointegration.

Investigation of ultrashort pulse laser ablation of the cornea and hydrogels for eye microsurgery

NASA Astrophysics Data System (ADS)

Girard, Guillaume; Zhou, Sheng; Bigaouette, Nicolas; Brunette, Isabelle; Chaker, Mohamed; Germain, Lucie; Lavertu, Pierre-Luc; Martin, François; Olivié, Gilles; Ozaki, Tsuneyuki; Parent, Mireille; Vidal, François; Kieffer, Jean-Claude

2004-10-01

The Femtosecond laser is a very promising tool for performing accurate dissection in various cornea layers. Clearly, the development of this application requires basic knowledge about laser-tissue interaction. One of the most significant parameter in laser applications is the ablation threshold, defined as the minimal laser energy per unit surface required for ablation. This paper investigates the ablation threshold as a function of the laser pulse duration for two corneal layers (endothelium and epithelium) as well as for hydrogel with different hydration degrees. The measured ablation thresholds prove to behave very differently as a function of the pulse duration for the various materials investigated, although the values obtained for the shortest laser pulses are quite similar. Our experimental results are fitted with a simple model for laser-matter interaction in order to determine some intrinsic physical parameters characterizing each target.

Femtosecond Laser Micro-structuring Of Transparent Materials And Its Ophthalmologic Applications

NASA Astrophysics Data System (ADS)

Ionin, Andrey A.; Kozhushko, Svetlana E.; Kudryashov, Sergey I.; Seleznev, Leonid V.; Sinitsyn, Dmitry V.; Zakharov, Stanislav D.; Alekhin, Alexander I.; Kourylyova, Irina M.; Kuz'min, Kirill K.; Likhvantseva, Vera G.; Samoylov, Michail V.

2010-10-01

We have successfully fabricated in vitro femtosecond laser micro-incisions inside cornea and—for the first time—inside sclera mildly pre-cleared by a biocompatible and clinically safe (non-toxic) natural agent (replacive refractive index-matching 40%-glucose solution in water), with the tissues taken as fresh cool cuts of human cadaver eyes, and reported on basic operational conditions of the micro-surgical procedures.

Femtosecond irradiation of chicken corneas analyzed by digital holographic microscopy

NASA Astrophysics Data System (ADS)

Fimia, A.; Gomariz, M.; Murciano, A.; Acebal, P.; Madrigal, R.; Carretero, L.; Alió, J. L.; Rodriguez, A.; Fernández, E.

2012-06-01

Digital Holographic Microscopy (DHM) is a potentially non-invasive new technology which can be applied in many areas from applied imaging science to biomedical optics. DHM is an interferometric technique that gives us a number of important advantages such as the possibility to acquire holograms at high speed, to obtain complete information about amplitude and phase and to use image processing techniques. In this sense, DHM offers rapid 3D imaging with a theoretically higher resolution than OCT (Optical Coherent Tomography). By this technique optical path measurements with sensitivities in the nanometer range of reflective and transparent objects can be obtained. In this work, we use DHM to study the effect of ablation using 4.5 nJ pulses on chicken corneas. For this, a titanium sapphire laser at 800 nm and 76 MHz frequency (Vitesse, Coherent Inc. USA) was focused to its diffraction-limited spot size by a 10x objective of 0.3 numerical aperture. The width of the pulse (170 fs) at the sample was measured by spectral techniques. The average beam power at the sample was 340 mW and all the system was mechanically driven by a XY synchronization unit that controls the speed of the sample movement. The speed of the sample was varied between 1-50 μm/s. The studied chicken corneal tissue was previously processed by Trypan dye in order to visualize the irradiated area. The photodisrupted zone was analyzed by a HDM technique by illuminating it using a laser diode source (λ=683 nm) linearly polarized in a modified Mach-Zehnder with an off-axis geometry configuration. The reflected object wave by the tissue surface (specimen) interferes with the reference wave and a CCD camera records the hologram. As a result, the influence of the speed of photodisruption in the depth of the ablated corneas was analyzed. Therefore, it is possible to analyze thermal and photoirradiated effects on corneal tissues which allow us the possibility to optimize the interaction of intratissue and the intratissue target region of interest.

The hierarchical response of human corneal collagen to load.

PubMed

Bell, J S; Hayes, S; Whitford, C; Sanchez-Weatherby, J; Shebanova, O; Vergari, C; Winlove, C P; Terrill, N; Sorensen, T; Elsheikh, A; Meek, K M

2018-01-01

Fibrillar collagen in the human cornea is integral to its function as a transparent lens of precise curvature, and its arrangement is now well-characterised in the literature. While there has been considerable effort to incorporate fibrillar architecture into mechanical models of the cornea, the mechanical response of corneal collagen to small applied loads is not well understood. In this study the fibrillar and molecular response to tensile load was quantified using small and wide angle X-ray scattering (SAXS/WAXS), and digital image correlation (DIC) photography was used to calculate the local strain field that gave rise to the hierarchical changes. A molecular scattering model was used to calculate the tropocollagen tilt relative to the fibril axis and changes associated with applied strain. Changes were measured in the D-period, molecular tilt and the orientation and spacing of the fibrillar and molecular networks. These measurements were summarised into hierarchical deformation mechanisms, which were found to contribute at varying strains. The change in molecular tilt is indicative of a sub-fibrillar "spring-like" deformation mechanism, which was found to account for most of the applied strain under physiological and near-physiological loads. This deformation mechanism may play an important functional role in tissues rich in fibrils of high helical tilt, such as skin and cartilage. Collagen is the primary mediator of soft tissue biomechanics, and variations in its hierarchical structure convey the varying amounts of structural support necessary for organs to function normally. Here we have examined the structural response of corneal collagen to tensile load using X-rays to probe hierarchies ranging from molecular to fibrillar. We found a previously unreported deformation mechanism whereby molecules, which are helically arranged relative to the axis of their fibril, change in tilt akin to the manner in which a spring stretches. This "spring-like" mechanism accounts for a significant portion of the applied deformation at low strains (<3%). These findings will inform the future design of collagen-based artificial corneas being developed to address world-wide shortages of corneal donor tissue. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Micromechanical Modeling Study of Mechanical Inhibition of Enzymatic Degradation of Collagen Tissues

PubMed Central

Tonge, Theresa K.; Ruberti, Jeffrey W.; Nguyen, Thao D.

2015-01-01

This study investigates how the collagen fiber structure influences the enzymatic degradation of collagen tissues. We developed a micromechanical model of a fibrous collagen tissue undergoing enzymatic degradation based on two central hypotheses. The collagen fibers are crimped in the undeformed configuration. Enzymatic degradation is an energy activated process and the activation energy is increased by the axial strain energy density of the fiber. We determined the intrinsic degradation rate and characteristic energy for mechanical inhibition from fibril-level degradation experiments and applied the parameters to predict the effect of the crimped fiber structure and fiber properties on the degradation of bovine cornea and pericardium tissues under controlled tension. We then applied the model to examine the effect of the tissue stress state on the rate of tissue degradation and the anisotropic fiber structures that developed from enzymatic degradation. PMID:26682825

Collagen Fibrils and Proteoglycans of Macular Dystrophy Cornea: Ultrastructure and 3D Transmission Electron Tomography.

PubMed

Akhtar, Saeed; Alkatan, Hind M; Kirat, Omar; Khan, Adnan A; Almubrad, Turki

2015-06-01

We report the ultrastructure and 3D transmission electron tomography of collagen fibrils (CFs), proteoglycans (PGs), and microfibrils within the CF of corneas of patients with macular corneal dystrophy (MCD). Three normal corneas and three MCD corneas from three Saudi patients (aged 25, 31, and 49 years, respectively) were used for this study. The corneas were processed for light and electron microscopy studies. 3D images were composed from a set of 120 ultrastructural images using the program "Composer" and visualized using the program "Visuliser Kai". 3D image analysis of MCD cornea showed a clear organization of PGs around the CF at very high magnification and degeneration of the microfibrils within the CF. Within the MCD cornea, the PG area in the anterior stroma was significantly larger than in the middle and posterior stroma. The PG area in the MCD cornea was significantly larger compared with the PG area in the normal cornea. The CF diameter and inter-fibrillar spacing of the MCD cornea were significantly smaller compared with those of the normal cornea. Ultrastructural 3D imaging showed that the production of unsulfated keratin sulfate (KS) may lead to the degeneration of micro-CFs within the CFs. The effect of the unsulfated KS was higher in the anterior stroma compared with the posterior stroma.

Anti-angiogenic potential of VEGF blocker dendron loaded on to gellan gum hydrogels for tissue engineering applications.

PubMed

Perugini, Valeria; Guildford, Anna L; Silva-Correia, Joana; Oliveira, Joaquim M; Meikle, Steven T; Reis, Rui L; Santin, Matteo

2018-02-01

Damage of non-vascularised tissues such as cartilage and cornea can result in healing processes accompanied by a non-physiological angiogenesis. Peptidic aptamers have recently been reported to block the vascular endothelial growth factor (VEGF). However, the therapeutic applications of these aptamers are limited due to their short half-life in vivo. In this work, an enhanced stability and bioavailability of a known VEGF blocker aptamer sequence (WHLPFKC) was pursued through its tethering of molecular scaffolds based on hyperbranched peptides, the poly(ɛ-lysine) dendrons, bearing three branching generations. The proposed design allowed simultaneous and orderly-spaced exposure of 16 aptamers per dendrimer to the surrounding biological microenvironent, as well as a relatively hydrophobic core based on di-phenylalanine aiming to promote an hydrophobic interaction with the hydrophobic moieties of ionically crosslinked methacrylated gellan gum (iGG-MA) hydrogels. The VEGF blocker dendrons were entrapped in iGG-MA hydrogels, and their capacity to prevent endothelial cell sprouting was assessed qualitatively and quantitatively using 3D in vitro models and the in vivo chick chorioallantoic membrane assay. The data demonstrate that at nanoscale concentrations, the dendronised structures were able to enhance control of the biological actvity of WHLPFKC at the material/tissue interface and hence the anti-angiogenic capacity of iGG-MA hydrogels not only preventing blood vessel invasion, but also inducing their regression at the tissue/iGG-MA interface. The in ovo study confirmed that iGG-MA functionalised with the dendron VEGF blockers do inhibit angiogenesis by controlling both size and ramifications of blood vessels in the proximity of the implanted gel surface. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Ultra-high speed OCT allows measurement of intraocular pressure, corneal geometry, and corneal stiffness using a single instrument

NASA Astrophysics Data System (ADS)

Singh, Manmohan; Han, Zhaolong; Nair, Achuth; Schill, Alexander; Twa, Michael D.; Larin, Kirill V.

2018-02-01

Screening for ocular diseases, such as glaucoma and keratoconus, includes measuring the eye-globe intraocular pressure (IOP) and corneal biomechanical properties. However, currently available clinical tools cannot quantify corneal tissue material parameters, which can provide critical information for detecting diseases and evaluating therapeutic outcomes. Here, we demonstrate measurement of eye-globe IOP, corneal elasticity, and corneal geometry of in situ porcine corneas with a technique termed applanation optical coherence elastography (Appl-OCE) with single instrument. We utilize an ultrafast phase-sensitive optical coherence tomography system comprised of a 4X buffered Fourier domain mode-locked swept source laser with an Ascan rate of 1.5 MHz and a 7.3 kHz resonant scanner. The IOP was measured by imaging the response of in situ porcine corneas to a large force air-puff. As with other noncontact tonometers, the time when the cornea was applanated during the inwards and outwards motion was correlated to a measure air-pressure temporal profile. The IOP was also measured with a commercially available rebound tonometer for comparison. The stiffness of the corneas was assessed by directly imaging and analyzing the propagation of a focused micro air-pulse induced elastic wave, and the corneal geometry was obtained from the OCT structural image. Our results show that corneal thickness decreased as IOP increased, and that corneal stiffness increased with IOP. Moreover, the IOP measurements made by Appl-OCE were more closely correlated with the artificially set IOP than the rebound tonometer, demonstrating the capabilities of Appl-OCE to measure corneal stiffness, eye-globe IOP, and corneal geometry with a single instrument.

Measurements of donor endothelial keratoplasty lenticules prepared from fresh donated whole eyes by using ultrasound and optical coherence tomography.

PubMed

Kanavi, Mozhgan Rezaei; Nemati, Farzan; Chamani, Tahereh; Kheiri, Bahar; Javadi, Mohammad Ali

2017-03-01

This study was conducted to analyze the profile and thickness of endothelial keratoplasty lenticules prepared from fresh donated whole eyes with Visante optical coherence tomography (V-OCT) compared to measurements obtained from ultrasound pachymetry (USP) at the Central Eye Bank of Iran. Microkeratome-assisted precut corneas were prepared for Descemet stripping automated endothelial keratoplasty by using standard eye bank protocol. Central posterior lenticule thickness (CPLT) on fresh whole eye, before excising corneoscleral disc and transferring to Optisol-GS, was measured by USP. V-OCT was used to measure central, paracentral, and midperipheral thicknesses of lenticules after transferring the tissue to Optisol-GS. Chi Square and Bonferroni tests were respectively used to uncover the differences between the USP and V-OCT measurements and also the thickness profile of lenticules. Postoperative reports for the entire transplanted lenticules were recorded. Accordingly, on evaluation of 312 enrolled precut corneas, CPLT measurements by V-OCT versus USP were statistically different (mean: 136 µm vs 165 µm, respectively; P = 0.008). Thickness profile of the posterior lenticules revealed increased thickness from the central to the peripheral parts of the cornea (mean increase of 16 µm at the pericentral and 64.2 µm at the peripheral locations, respectively); however, the increase in the thickness was relatively symmetrical. Postoperative reports of transplanted lenticules were unremarkable, since there were no posterior flap detachments. In essence, V-OCT measurements of microkeratome-assisted precut lenticules prepared from fresh donated whole eyes averaged 29 μm thinner than USP measurements and revealed a significant but symmetric increase of thickness towards the peripheral parts of the corneas. However, the variation in the thickness profile did not affect the attachment or the clarity of transplanted precut lenticlues.

An Evaluation of Lysyl Oxidase–Derived Cross-Linking in Keratoconus by Liquid Chromatography/Mass Spectrometry

PubMed Central

Takaoka, Anna; Babar, Natasha; Hogan, Julia; Kim, MiJung; Price, Marianne O.; Price, Francis W.; Trokel, Stephen L.; Paik, David C.

2016-01-01

Purpose Current literature contains scant information regarding the extent of enzymatic collagen cross-linking in the keratoconus (KC) cornea. The aim of the present study was to examine levels of enzymatic lysyl oxidase–derived cross-links in stromal collagen in KC tissue, and to correlate the cross-link levels with collagen fibril stability as determined by thermal denaturation temperature (Tm). Methods Surgical KC samples (n = 17) and Eye-Bank control (n = 11) corneas of age 18 to 68 years were analyzed. The samples were defatted, reduced (NaBH4), hydrolyzed (6N HCl at 110°C for 18 hours), and cellulose enriched before analysis by C8 high-performance liquid chromatography equipped with parallel fluorescent and mass detectors in selective ion monitoring mode (20 mM heptafluorobutyric acid/methanol 70:30 isocratic at 1 mL/min). Nine different cross-links were measured, and the cross-link density was determined relative to collagen content (determined colorimetrically). The Tm was determined by differential scanning calorimetry. Results Cross-links detected were dihydroxylysinonorleucine (DHLNL), hydroxylysinonorleucine, lysinonorleucine (LNL), and histidinohydroxylysinonorleucine in both control and KC samples. Higher DHLNL levels were detected in KC, whereas the dominant cross-link, LNL, was decreased in KC samples. Decreased LNL levels were observed among KC ≤ 40 corneas. There was no difference in total cross-link density between KC samples and the controls. Pyridinolines, desmosines, and pentosidine were not detected. There was no notable correlation between cross-link levels with fibril instability as determined by Tm. Conclusions Lower levels of LNL in the KC cornea suggest that there might be a cross-linking defect either in fibrillar collagen or the microfibrillar elastic network composed of fibrillin. PMID:26780316

DenTimol as A Dendrimeric Timolol Analogue for Glaucoma Therapy: Synthesis and Preliminary Efficacy and Safety Assessment.

PubMed

Lancina, Michael G; Wang, Juan; Williamson, Geoffrey S; Yang, Hu

2018-05-30

In this work, we report the synthesis and characterization of DenTimol, a dendrimer-based polymeric timolol analog, as a glaucoma medication. A timolol precursor ( S)-4-[4-(oxiranylmethoxy)-1,2,5-thiadiazol-3-yl]morpholine (OTM) was reacted with the heterobifunctional amine polyethylene glycol acetic acid (amine-PEG-acetic acid, M n = 2000 g/mol) via a ring opening reaction of an epoxide by an amine to form the OTM-PEG conjugate. OTM-PEG was then coupled to an ethylenediamine (EDA) core polyamidoamine (PAMAM) dendrimer G3 to generate DenTimol using the N-(3-(dimethylamino)propyl)- N'-ethylcarbodiimide hydrochloride (EDC)/ N-hydroxysuccinimide (NHS) coupling reaction. MALDI mass spectrometry, 1 H NMR spectroscopy, and HPLC were applied to characterize the intermediate and final products. Ex vivo corneal permeation of DenTimol was assessed using the Franz diffusion cell system mounted with freshly extracted rabbit cornea. The cytotoxicity of DenTimol was assessed using the WST-1 assay. Our results show that DenTimol is nontoxic up to an OTM equivalent concentration of 100 μM. DenTimol is efficient at crossing the cornea. About 8% of the dendrimeric drug permeated through the cornea in 4 h. Its IOP-lowering effect was observed in normotensive adult Brown Norway male rats. Compared to the undosed eye, an IOP reduction by an average of 7.3 mmHg (∼30% reduction from baseline) was observed in the eye topically treated with DenTimol (2 × 5 μL, 0.5% w/v timolol equivalent) in less than 30 min. Daily dosing of DenTimol for a week did not cause any irritation or toxicity as confirmed by the histological examination of ocular tissues, including the cornea, ciliary body, and retina.

Developing an in situ nanosuspension: a novel approach towards the efficient administration of poorly soluble drugs at the anterior eye.

PubMed

Luschmann, Christoph; Tessmar, Joerg; Schoeberl, Simon; Strauss, Olaf; Framme, Carsten; Luschmann, Karl; Goepferich, Achim

2013-11-20

With about 50-60 million cases in the US alone, dry eye disease represents a severe health care problem. Cyclosporin A (CsA) would be a potent candidate for a causal therapy. However, CsA is not sufficiently water soluble to be administrated via simple eye drops. We developed an in situ nanosuspension (INS) as a novel approach towards the administration of CsA to the cornea. It precipitates upon contact with the tear fluid and creates CsA nanoparticles that enter the cornea and release the drug by dissolution. We selected two liquid poly(ethylene glycols) (PEG) that dissolve CsA and create nanoparticles by precipitation of CsA upon water contact. Aqueous solutions of PEG and Solutol, a non-ionic surfactant, were well tolerated by primary human epithelial cells in vitro. To determine the critical water content needed for a precipitation, the solubility of CsA was investigated in quaternary systems of drug, solvent, surfactant and water. The best INS formulation showed a particle size of 505 ± 5 nm, a polydispersity index (PdI) of 0.23 ± 0.03 and a neutral zeta potential of -0.07 ± 0.05 mV. After single administration to porcine eyes in vitro, 3165 ± 597 ng(CsA)/g(cornea) were detected in corneal tissue, while the levels of Restasis a commercial formulation were, with 545 ± 137 ng(CsA)/g(cornea), significantly lower (P<0.01). These results demonstrate that an INS is a promising, novel approach towards the causal treatment of inflammatory diseases at the anterior eye. Copyright © 2013 Elsevier B.V. All rights reserved.

Influence of Pre-Freezing Temperature on the Corneal Endothelial Cytocompatibility and Cell Delivery Performance of Porous Hyaluronic Acid Hydrogel Carriers.

PubMed

Lai, Jui-Yang

2015-08-11

The development of porous hyaluronic acid (HA) hydrogels for corneal endothelial tissue engineering is attractive because they can be used as functional cell delivery carriers to help in the reconstruction of damaged areas. The purpose of this study was to investigate the corneal endothelial cytocompatibility and cell delivery performance of porous HA hydrogel biomaterials fabricated at different pre-freezing temperatures. As compared to their counterparts prepared at -80 °C, the HA samples fabricated at higher pre-freezing temperature (i.e., 0 °C) exhibited a larger pore size and higher porosity, thereby leading to lower resistance to glucose permeation. Live/dead assays and gene expression analyses showed that the restricted porous structure of HA carriers decreases the viability and ionic pump function of cultured corneal endothelial cells (CECs). The results also indicated that the porous hydrogel biomaterials fabricated at high pre-freezing temperature seem to be more compatible with rabbit CECs. In an animal model of corneal endothelial dysfunction, the wounded rabbit corneas receiving bioengineered CEC sheets and restricted porous-structured HA carriers demonstrated poor tissue reconstruction. The therapeutic efficacy of cell sheet transplants can be improved by using carrier materials prepared at high pre-freezing temperature. Our findings suggest that the cryogenic operation temperature-mediated pore microstructure of HA carriers plays an important role in corneal endothelial cytocompatibility and cell delivery performance.

The use of topical honey in the treatment of corneal abrasions and endotoxin-induced keratitis in an animal model.

PubMed

Uwaydat, Sami; Jha, Purushottam; Tytarenko, Ruslana; Brown, Harry; Wiggins, Michael; Bora, Puran S; Bora, Nalini S

2011-09-01

 To investigate the effect of topically applied honey on intact corneas, surgically induced corneal abrasions and endotoxin induced keratitis. The effect of honey on the cornea was investigated by application of honey on intact corneas, wounded corneas and endotoxin-induced keratitis in Lewis rats. The corneas were wounded by creating an epithelial defect using a surgical blade, and the keratitis was induced by topically applying Pseudomonas aeruginosa endotoxin to scarified corneas. After treatment rats were sacrificed and cornea harvested in each case. Corneas were processed for paraffin embedding for histological and immuno-fluorescence staining. Corneas were also harvested and processed for total ribonucleic acid (RNA) isolation for reverse transcriptase-polymerase chain reaction (RT-PCR) analysis for various growth factors and inflammatory chemokines/cytokines).  Histological analysis revealed that no inflammation or morphological changes occurred after honey treatment in naive intact corneas. Vascular endothelial growth factor (VEGF) levels were also not altered after honey treatment. Topical application of honey to injured corneas resulted in faster epithelial healing and decreased expression of VEGF, transforming growth factor beta (TGF-β), interferon gamma (IFN-γ), interleukin 12 (IL-12) and tumor necrosis factor alpha (TNF-α) in injured corneas. Our results also established that honey treatment reduced the inflammation in endotoxin-induced keratitis by reducing the levels of angiogenic factors (VEGF and TGF-β), inflammatory cytokines (IL-12) and chemokines (CC chemokine receptor 5(CCR-5)).  Short term use of honey on intact corneas can be safe. Honey has anti-angiogenic and anti-inflammatory properties that can be explored in several corneal inflammatory and infectious conditions.

Olfactomedin-like 2 A and B (OLFML2A and OLFML2B) expression profile in primates (human and baboon).

PubMed

Pérez-Ibave, Diana Cristina; González-Alvarez, Rafael; de La Luz Martinez-Fierro, Margarita; Ruiz-Ayma, Gabriel; Luna-Muñoz, Maricela; Martínez-De-Villarreal, Laura Elia; De Lourdes Garza-Rodríguez, María; Reséndez-Pérez, Diana; Mohamed-Noriega, Jibran; Garza-Guajardo, Raquel; Bautista-De-Lucío, Víctor Manuel; Mohamed-Noriega, Karim; Barboza-Quintana, Oralia; Arámburo-De-La-Hoz, Carlos; Barrera-Saldaña, Hugo Alberto; Rodríguez-Sánchez, Irám Pablo

2016-11-08

The olfactomedin-like domain (OLFML) is present in at least four families of proteins, including OLFML2A and OLFML2B, which are expressed in adult rat retina cells. However, no expression of their orthologous has ever been reported in human and baboon. The aim of this study was to investigate the expression of OLFML2A and OLFML2B in ocular tissues of baboons (Papio hamadryas) and humans, as a key to elucidate OLFML function in eye physiology. OLFML2A and OLFML2B cDNA detection in ocular tissues of these species was performed by RT-PCR. The amplicons were cloned and sequenced, phylogenetically analyzed and their proteins products were confirmed by immunofluorescence assays. OLFML2A and OLFML2B transcripts were found in human cornea, lens and retina and in baboon cornea, lens, iris and retina. The baboon OLFML2A and OLFML2B ORF sequences have 96% similarity with their human's orthologous. OLFML2A and OLFML2B evolution fits the hypothesis of purifying selection. Phylogenetic analysis shows clear orthology in OLFML2A genes, while OLFML2B orthology is not clear. Expression of OLFML2A and OLFML2B in human and baboon ocular tissues, including their high similarity, make the baboon a powerful model to deduce the physiological and/or metabolic function of these proteins in the eye.

Variations in the endogenous fluorescence of rabbit corneas after mechanical property alterations

NASA Astrophysics Data System (ADS)

Ortega-Martinez, Antonio; Touchette, Genna; Zhu, Hong; Kochevar, Irene E.; Franco, Walfre

2017-09-01

Keratoconus is an eye disease in which the cornea progressively deforms due to loss of cornea mechanical rigidity, and thus causes deterioration of visual acuity. Techniques to characterize the mechanical characteristics of the cornea are important to better monitor changes and response to treatments. To investigate the feasibility of using the endogenous fluorescence of cornea for monitoring alterations of its mechanical rigidity, linear tensiometry was used to quantitate stiffness and Young's modulus (YM) after treatments that increase cornea stiffness (collagen photocross-linking) or decrease stiffness (enzymatic digestion). The endogenous ultraviolet fluorescence of cornea was also measured before and after these treatments. The fluorescence excitation/emission spectral ranges were 280 to 430/390 to 520 nm, respectively. A correlation analysis was carried out to identify fluorescence excitation/emission pairs whose intensity changes correlated with the stiffness. A positive correlation was found between variations in fluorescence intensity of the 415-/485-nm excitation/emission pair and YM of photocross-linked corneas. After treatment of corneas with pepsin, the YM decreased as the fluorescence intensity at 290-/390-nm wavelengths decreased. For weakening of corneas with collagenase, only qualitative changes in the fluorescence spectrum were observed. Changes in the concentration of native or newly created fluorescent molecular species contain information that may be directly or indirectly related to the mechanical structure of the cornea.

Cryopreservation of rabbit corneas: assessment by microscopy and transplantation.

PubMed Central

Fong, L P; Hunt, C J; Taylor, M J; Pegg, D E

1986-01-01

Rabbit corneas were frozen and thawed by three methods and compared by full thickness transplantation as well as specular microscopy, histology, and transmission electron microscopy. Two of the methods used a recently described technique, in which the excised cornea was immersed in a potassium-rich buffered solution containing the cryoprotectant dimethyl sulphoxide (Me2SO, 2 mol/l). This solution was designed to restrict the loss of intracellular potassium and to prevent cell swelling at low temperatures. In one group the corneas were frozen and thawed surrounded by 5 ml of medium, while in the second group corneas were drained of excess fluid and frozen in air. The third group consisted of corneas cryopreserved by Capella and colleagues' method. All the cryopreserved corneas were damaged, but those that had been frozen in air after exposure to the new medium showed better structure and function than corneas frozen by either of the other two techniques. Images PMID:3535871

The structure of interfibrillar proteoglycan bridges (‘shape modules’) in extracellular matrix of fibrous connective tissues and their stability in various chemical environments

PubMed Central

SCOTT, JOHN E.; THOMLINSON, ALISON M.

1998-01-01

Collagen fibrils in extracellular matrices of connective tissues (tendon, cornea, etc.) are bridged and linked by the anionic glycosaminoglycans (AGAGs) of the small proteoglycans (decoron, etc.). It was proposed that these bridges and ties maintain the collagen fibril dispositions in relation to each other, helping to define tissue shape, and hence called shape modules. This investigation describes chemical and physicochemical conditions in which these structures are stable and what treatments cause their disruption. The effects on fixed and unfixed sections of tendon, cornea, lung and ear from rat, mouse and rabbit of pH, electrolyte concentration, EDTA, mercaptoethanol, hydrogen peroxide, free radicals, periodate, acetylation, urea, nonionic detergent and organic solvents were assessed by staining with Cupromeronic blue or Alcec blue in CEC techniques to localise AGAG bridges or their disintegration products. Ca2+ was not involved in the structures, oxidation/reduction had no effect and Triton X100, a nonionic detergent did not damage them. They were stable between pH 4.5 and 9.5. Periodate as a glycol-cleaving reagent did not affect them. High concentrations of urea (>2.0 m) and MgCl2 (0.5 m) disrupted the tissues. The combination of Triton and urea at concentrations too low to cause damage separately was disruptive. Free radicals in periodate solutions were damaging. Organic solvents caused collapse and rearrangements of the AGAG filaments. Acetylation caused considerable disruption of shape modules. Dermochondan but not keratan sulphate AGAGs were removed by treatment with NaOH. After fixing with glutaraldehyde only free radical and NaOH treatments were severely disruptive of shape modules. The results are compatible with a previously proposed structure for the shape modules, stabilised by hydrophobic and hydrogen bonding. PMID:9688505

Vision Restoration with a Collagen Crosslinked Boston Keratoprosthesis Unit

DTIC Science & Technology

2016-09-01

AWARD NUMBER: W81XWH-15-2-0044 TITLE: Vision Restoration with a Collagen Crosslinked Boston Keratoprosthesis Unit PRINCIPAL INVESTIGATOR: Joseph B...TITLE AND SUBTITLE 5a. CONTRACT NUMBER Vision Restoration with a Collagen Cross-linked Boston Keratoprosthesis Unit 5b. GRANT NUMBER W81XWH-15-2-0044...the incidence or potentially eliminate corneal melts by strengthening the keratoprosthesis carrier tissue by collagen -crosslinking the cornea graft

Effectiveness of a decontamination method for donor corneas.

PubMed

Badenoch, P R; Alfrich, S J; Wedding, T R; Coster, D J

1988-03-01

A retrospective study was made of the effectiveness of an eye bank decontamination and storage method. A comparison was made between microbial cultures taken from the limbus at enucleation and from scleral remnants recovered after surgery. Organisms were isolated from the limbus of 73% of donor eyes and from 4% of remnants. Standard eye bank procedures were found to eradicate gut and skin organisms, including candida, from donor tissue.

A Class I (Senofilcon A) Soft Contact Lens Prevents UVB-Induced Ocular Effects, Including Cataract, in the Rabbit In Vivo

PubMed Central

Lin, Li-Ren; Leverenz, Victor R.; Dang, Loan

2011-01-01

Purpose. UVB radiation from sunlight is known to be a risk factor for human cataract. The purpose in this study was to investigate the ability of a class I UV-blocking soft contact lens to protect against UVB-induced effects on the ocular tissues of the rabbit in vivo. Methods. Eyes of rabbits were exposed to UVB light for 30 minutes (270–360 nm, peak at 310 nm, 1.7 mW/cm2 on the cornea). Eyes were irradiated in the presence of either a UV-blocking senofilcon A contact lens, a minimally UV-blocking lotrafilcon A contact lens, or no contact lens at all. Effects on the cornea and lens were evaluated at various times after exposure. Results. Eyes irradiated with no contact lens protection showed corneal epithelial cell loss plus lens epithelial cell swelling, vacuole formation, and DNA single-strand breaks, as well as lens anterior subcapsular opacification. The senofilcon A lens protected nearly completely against the UVB-induced effects, whereas the lotrafilcon A lens showed no protection. Conclusions. The results indicate that use of a senofilcon A contact lens is beneficial in protecting ocular tissues of the rabbit against the harmful effects of UVB light, including photokeratitis and cataract. PMID:21421866

A class I (Senofilcon A) soft contact lens prevents UVB-induced ocular effects, including cataract, in the rabbit in vivo.

PubMed

Giblin, Frank J; Lin, Li-Ren; Leverenz, Victor R; Dang, Loan

2011-06-01

UVB radiation from sunlight is known to be a risk factor for human cataract. The purpose in this study was to investigate the ability of a class I UV-blocking soft contact lens to protect against UVB-induced effects on the ocular tissues of the rabbit in vivo. Eyes of rabbits were exposed to UVB light for 30 minutes (270-360 nm, peak at 310 nm, 1.7 mW/cm(2) on the cornea). Eyes were irradiated in the presence of either a UV-blocking senofilcon A contact lens, a minimally UV-blocking lotrafilcon A contact lens, or no contact lens at all. Effects on the cornea and lens were evaluated at various times after exposure. Eyes irradiated with no contact lens protection showed corneal epithelial cell loss plus lens epithelial cell swelling, vacuole formation, and DNA single-strand breaks, as well as lens anterior subcapsular opacification. The senofilcon A lens protected nearly completely against the UVB-induced effects, whereas the lotrafilcon A lens showed no protection. The results indicate that use of a senofilcon A contact lens is beneficial in protecting ocular tissues of the rabbit against the harmful effects of UVB light, including photokeratitis and cataract.

Formulation Development, Optimization, and In vitro - In vivo Characterization of Natamycin Loaded PEGylated Nano-lipid Carriers for Ocular Applications.

PubMed

Patil, Akash; Lakhani, Prit; Taskar, Pranjal; Wu, Kai-Wei; Sweeney, Corinne; Avula, Bharathi; Wang, Yan-Hong; Khan, Ikhlas A; Majumdar, Soumyajit

2018-04-23

Current study aimed at formulating and optimizing natamycin (NT) loaded PEGylated NLCs (NT-PEG-NLCs) using Box-Behnken Design and investigating their potential in ocular applications. Response surface methodology (RSM) computations and plots for optimization were performed using Design Expert ® software, to obtain optimum values for response variables based on the criteria of desirability. Optimized NT-PEG-NLCs had predicted values for the dependent variables not significantly different from the experimental values. NT-PEG-NLCs were characterized for their physicochemical parameters; NT's rate of permeation and flux across rabbit cornea was evaluated, in vitro; ocular tissue distribution was assessed in rabbits, in vivo. NT-PEG-NLCs were found to have optimum particle size (< 300 nm) narrow PDI, high NT entrapment and NT content. In vitro transcorneal permeability and flux of NT from NT-PEG-NLCs was significantly higher than Natacyn ® . NT-PEG-NLC (0.3%) showed improved delivery of NT across the intact cornea and provided concentrations statistically similar to the marketed suspension (5%) in inner ocular tissues, in vivo, indicating that it could be a potential alternative to the conventional suspension during the course of fungal keratitis therapy. Copyright © 2018. Published by Elsevier Inc.

Decorin and biglycan of normal and pathologic human corneas

NASA Technical Reports Server (NTRS)

Funderburgh, J. L.; Hevelone, N. D.; Roth, M. R.; Funderburgh, M. L.; Rodrigues, M. R.; Nirankari, V. S.; Conrad, G. W.

1998-01-01

PURPOSE: Corneas with scars and certain chronic pathologic conditions contain highly sulfated dermatan sulfate, but little is known of the core proteins that carry these atypical glycosaminoglycans. In this study the proteoglycan proteins attached to dermatan sulfate in normal and pathologic human corneas were examined to identify primary genes involved in the pathobiology of corneal scarring. METHODS: Proteoglycans from human corneas with chronic edema, bullous keratopathy, and keratoconus and from normal corneas were analyzed using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), quantitative immunoblotting, and immunohistology with peptide antibodies to decorin and biglycan. RESULTS: Proteoglycans from pathologic corneas exhibit increased size heterogeneity and binding of the cationic dye alcian blue compared with those in normal corneas. Decorin and biglycan extracted from normal and diseased corneas exhibited similar molecular size distribution patterns. In approximately half of the pathologic corneas, the level of biglycan was elevated an average of seven times above normal, and decorin was elevated approximately three times above normal. The increases were associated with highly charged molecular forms of decorin and biglycan, indicating modification of the proteins with dermatan sulfate chains of increased sulfation. Immunostaining of corneal sections showed an abnormal stromal localization of biglycan in pathologic corneas. CONCLUSIONS: The increased dermatan sulfate associated with chronic corneal pathologic conditions results from stromal accumulation of decorin and particularly of biglycan in the affected corneas. These proteins bear dermatan sulfate chains with increased sulfation compared with normal stromal proteoglycans.

Simulation model of an eyeball based on finite element analysis on a supercomputer.

PubMed

Uchio, E; Ohno, S; Kudoh, J; Aoki, K; Kisielewicz, L T

1999-10-01

A simulation model of the human eye was developed. It was applied to the determination of the physical and mechanical conditions of impacting foreign bodies causing intraocular foreign body (IOFB) injuries. Modules of the Hypermesh (Altair Engineering, Tokyo, Japan) were used for solid modelling, geometric construction, and finite element mesh creation based on information obtained from cadaver eyes. The simulations were solved by a supercomputer using the finite element analysis (FEA) program PAM-CRASH (Nihon ESI, Tokyo, Japan). It was assumed that rupture occurs at a strain of 18.0% in the cornea and 6.8% in the sclera and at a stress of 9.4 MPa for both cornea and sclera. Blunt-shaped missiles were shot and set to impact on the surface of the cornea or sclera at velocities of 30 and 60 m/s, respectively. According to the simulation, the sizes of missile above which corneal rupture occurred at velocities of 30 and 60 m/s were 1.95 and 0.82 mm. The missile sizes causing scleral rupture were 0.95 and 0.75 mm at velocities of 30 and 60 m/s. These results suggest that this FEA model has potential usefulness as a simulation tool for ocular injury and it may provide useful information for developing protective measures against industrial and traffic ocular injuries.

[Correlations of telomere length changing and pathogeny of keratoconus].

PubMed

Wang, Jiao-jiao; Li, Shao-wei; Wang, Yi-qiang; Wang, Ye; Zhong, Wen-xian; Zang, Xin-jie

2009-08-01

To study telomere length, senescence-associated-beta-galactosidase (SA-beta-galactosidase) and senescence marker protein-30 (SMP-30) in the stromal cells of keratoconus or normal corneas respectively, aiming finding the association of these indexes with the phenotype of keratoconus. Experiment research. 37 keratoconus lesions corneas were removed from 32 keratoconus patients who were operated in Shangdong Eye Institute between January 2006 and December 2006, and 20 normal corneas were collected from eye bank. The keratoconus corneas ages were from 13 to 34 years [mean ages (19 + or - 5) years] and the control group consists of 20 normal corneas donor ages from 9 to 30 years [mean ages (19 + or - 4) years]. And there was no statistical difference of ages between keratoconus and normal corneas. Southern blot method was utilized to detect telomere length of genomic DNA. SA-beta-galactosidase was detected by 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-Gal) staining method respectively in keratoconus and normal corneas. Isolated mRNA from keratoconus and normal corneas were reverse-transcribed to cDNA and SMP-30 was detected using PCR with specific primers (sense: 5' ccg tgg atg cct ttg act at 3'; anti-sense: 5' caa ctt cat gc tgct ttg ga 3'). To compare normal corneas and keratoconus corneas by histopathological study. Statistical analysis by t test. The telomere length in stromal cells in keratoconus corneas were from 10.29 to 14.12 kb, mean (11.54 + or - 1.41) kb, while that of normal corneas were from 12.64 to 15.32 kb, mean (13.45 + or - 0.99) kb. The difference of telomere length in stromal cells of keratoconus and normal corneas reached a statistical significant level (t = 4.753, P < 0.05). That means the telomere length of keratoconus stroma was shorter than that of normal corneal stroma. Light microscopy revealed that collagen fibers in keratoconus corneal stroma were arranged in an irregular manner. Cells density in keratoconus stroma appeared lower than in normal ones but the decrease was not significant. The staining of SA-beta-galactosidase in the keratoconus section was evident, but there was no staining in the normal corneas. SMP-30 was not detectable with RT-PCR method in either keratoconus or normal corneas. Telomeres in the keratoconus stromas manifest higher SA-beta-galactosidase than control, implying that improper senescence might be involved in pathogenesis of keratoconus.

Vector delivery technique affects gene transfer in the cornea in vivo.

PubMed

Mohan, Rajiv R; Sharma, Ajay; Cebulko, Tyler C; Tandon, Ashish

2010-11-27

This study tested whether controlled drying of the cornea increases vector absorption in mouse and rabbit corneas in vivo and human cornea ex vivo, and studied the effects of corneal drying on gene transfer, structure and inflammatory reaction in the mouse cornea in vivo. Female C57 black mice and New Zealand White rabbits were used for in vivo studies. Donor human corneas were used for ex vivo experiments. A hair dryer was used for drying the corneas after removing corneal epithelium by gentle scraping. The corneas received no, once, twice, thrice, or five times warm air for 10 s with a 5 s interval after each 10 s hair dryer application. Thereafter, balanced salt solution (BSS) was topically applied immediately on the cornea for 2 min using a custom-cloning cylinder. The absorbed BSS was quantified using Hamilton microsyringes. The adeno-associated virus 8 (AAV8) vector (1.1×10(8) genomic copies/µl) expressing marker gene was used to study the effect of corneal drying on gene transfer. Animals were sacrificed on day 14 and gene expression was analyzed using commercial staining kit. Morphological changes and infiltration of inflammatory cells were examined with H & E staining and immunocytochemistry. Mice, rabbit or human corneas subjected to no or 10 s drying showed 6%-8% BSS absorption whereas 20, 30, or 50 s corneal drying showed significantly high 14%-19% (p<0.001), 21%-22% (p<0.001), and 25%-27% (p<0.001) BSS absorption, respectively. The AAV8 application on mouse cornea after 50 s drying showed significantly higher transgene delivery (p<0.05) in vivo with mild-to-moderate changes in corneal morphology. The 30 s of drying also showed significantly (p<0.05) high transgene delivery in mouse stroma in vivo without jeopardizing corneal morphology whereas 10 or 20 s drying showed moderate degree of gene transfer with no altered corneal morphology. Corneas that underwent 50 s drying showed high CD11b-positive cells (p<0.01) compared to control corneas whereas 20 or 30 s air-dried corneas showed insignificant CD11b-positive cells compared to control corneas. Controlled corneal drying with hair dryer increases vector absorption significantly. The dispensing of efficacious AAV serotype into cornea with optimized minimally invasive topical application technique could provide high and targeted expression of therapeutic genes in the stroma in vivo without causing significant side effects.

Ultrastructure features of camel cornea--collagen fibril and proteoglycans.

PubMed

Almubrad, Turki; Akhtar, Saeed

2012-01-01

  The uniform distribution of collagen fibrils and proteoglycans maintain the transparency of normal cornea. We describe the ultrastructural features of camel cornea including collagen fibrils and proteoglycans (PGs).   Camel corneas (of 6-, 8-, and 10-month-old animals) were fixed in 2.5% glutaraldehyde containing cuprolinic blue in sodium acetate buffer and processed for electron microscopy. The 'AnalySIS LS Professional' program was used to analyze the collagen fibril diameter.   The camel cornea consists of four layers: the epithelium (227 μm), stroma (388 μm), Descemet's membrane (DM), and endothelium. The epithelium constituted 36% of the camel cornea, whereas corneal stroma constituted 62% of the corneal thickness (629 μm). The PGs in the posterior stroma were significantly larger in number and size compared with the anterior and middle stroma. The collagen fibril diameter was 25 nm and interfibrillar spacing 40 nm. Fibrillar structures are present throughout the DM.   The structure of the camel cornea is very different from human and other animals. The unique structure of the cornea might be an adaptation to help the camel to survive in a hot and dry climate. The camel cornea may also be a good model to study the effect of hot and dry climates on the cornea. © 2011 American College of Veterinary Ophthalmologists.

Protein structural failure in mid-IR laser ablation of cornea

NASA Astrophysics Data System (ADS)

Hutson, M. Shane; Xiao, Yaowu; Guo, Mingsheng

2006-05-01

Researchers have previously observed that tissue ablation with a free electron laser tuned to wavelengths between 6-7 μm is accompanied by remarkably little collateral damage. Attempts to explain these observations have invoked a wavelength-dependent loss of protein structural integrity; however, the molecular nature of this structural failure has been heretofore ill-defined. In this report, we evaluate several candidates for the relevant transition by analyzing the non-volatile debris ejected during ablation. Porcine corneas were ablated with a free electron laser tuned to either 2.77 or 6.45 μm - wavelengths that are equally well absorbed by hydrated corneas, but that respectively target water or protein as the primary chromophore. The ejected debris was characterized via gel electrophoresis, as well as FTIR, micro-Raman and 13C-NMR spectroscopy. We find that high-fluence (240 J/cm2) ablation at 6.45 μm, but not at 2.77 μm, leads to protein fragmentation. This fragmentation is accompanied by the accumulation of nitrile and alkyne species. Although these initial experiments did not detect significant protein unfolding, the loss of collagen triple-helix structure was evident using UV and vibrational circular dichroism. The candidate transition most consistent with all these observations is scission of the collagen protein backbone at N-alkylamide bonds. Identifying this transition is a key step towards understanding the observed wavelength-dependence of collateral damage.

New screening methods for donor eye-bank eyes.

PubMed

Terry, M A; Ousley, P J

1999-07-01

Current methods of screening donor eyes for corneal transplantation are not always effective in excluding corneas with abnormal topography. We used the Orbscan to determine whether corneal-thickness maps could be used as a technique for donor tissue screening. Forty eye-bank eyes were measured with the Orbscan, and a corneal-thickness map was generated. Average central pachymetry measurements from each map were compared with the thinnest midperipheral thickness reading. Two eyes from a donor who had photorefractive keratectomy (PRK) and two eyes from a donor with keratoconus were then compared with the normal donor eye results. The average difference between the thinnest midperipheral pachymetry and the central pachymetry in the control group was 0.040 +/- 0.026 mm. The eyes from the donor with PRK showed larger disparities between the central and midperipheral thicknesses because of the thinned central cornea, with differences of 0.154 mm in the right eye and 0.106 mm in the left eye. The eyes from the donor with keratoconus had midperipheral corneas that were thinner than the center, indicating eccentric, ectatic cones. The differences in thickness between the center and midperiphery in the eyes from the donor with PRK and the donor with keratoconus differed from the control group by >2 SD. Diseases or surgery that affect the relationship between the central and midperipheral corneal thickness may be screened through Orbscan pachymetry mapping with comparison with a normal range.

Corneal confocal microscopy following conventional, transepithelial, and accelerated corneal collagen cross-linking procedures for keratoconus.

PubMed

Touboul, David; Efron, Nathan; Smadja, David; Praud, Delphine; Malet, Florence; Colin, Joseph

2012-11-01

To compare early corneal healing following conventional, transepithelial, and accelerated corneal collagen cross-linking (CXL) protocols. Twenty-four patients with progressive keratoconus were divided into three groups to receive conventional, transepithelial, or accelerated CXL. In vivo corneal confocal microscopy was performed on each patient preoperatively and at 1, 3, and 6 months postoperatively. Closure of the epithelial wound was complete 3 days following conventional and accelerated CXL. The subbasal nerve plexus was essentially obliterated immediately following conventional and accelerated CXL, and virtually no nerve fibers had regenerated by 6 months. The anterior stroma showed significant changes 1 month following conventional CXL; these changes were similar but more pronounced following accelerated CXL. Observed stromal changes included complete obliteration of keratocytes, increased tissue reflectivity, a honeycomb-like appearance, and circular lacunae. Some recovery of keratocyte density was noted after 6 months. These changes were less pronounced in the mid-stroma, and there were no apparent changes to the posterior stroma or endothelium. The cornea appeared to be unaltered following transepithelial CXL. In vivo corneal confocal microscopy analysis of the postoperative impact of CXL on the cornea revealed clear differences among conventional, accelerated, and transepithelial CXL protocols. Accelerated CXL had a greater impact than conventional CXL on the anterior cornea, whereas transepithelial CXL did not appear to alter corneal morphology. Copyright 2012, SLACK Incorporated.

A novel explanation of corneal clouding in a bone marrow transplant-treated patient with Hurler syndrome

PubMed Central

Yuan, Ching; Bothun, Erick D.; Hardten, David R.; Tolar, Jakub; McLoon, Linda K.

2016-01-01

One common complication of mucopolysaccharidosis I-Hurler (MPS1-H) is corneal clouding, which occurs despite current treatments, including bone marrow transplantation. Human corneas were obtained from a 14 year old subject with MPS1-H and visual disability from progressive corneal clouding despite a prior bone marrow transplant at age 2. This was compared to a cornea from a 17 year old donated to our eye bank after his accidental death. The corneas were analyzed microscopically after staining with Alcian blue, antibodies to collagen I, IV, VI, and α-smooth muscle actin. Differences in levels of expression of the indicated molecules were assessed. Corneas from Hurler and control mice were examined similarly to determine potential mechanistic overlap. The MPS1-H subject cornea showed elevations in mucopolysaccharide deposition. The MPS1-H and Hurler mice corneas showed increased and disorganized expression of collagen I and IV relative to the control corneas. The MPS1-H corneas also showed increased and disordered expression of collagen VI. Positive expression of α-smooth muscle actin indicated myofibroblast conversion within the MPS1-H cornea in both the patient and mutant mouse material compared to normal human and control mouse cornea. Increased deposition of collagens and smooth muscle actin correlate with corneal clouding, providing a potential mechanism for corneal clouding despite bone marrow transplantation in MPS1-H patients. It might be possible to prevent or slow the onset of corneal clouding by treating the cornea with drugs known to prevent myofibroblast conversion. PMID:27235795

Limbal Stem Cell Deficiency and Treatment with Stem Cell Transplantation.

PubMed

Barut Selver, Özlem; Yağcı, Ayşe; Eğrilmez, Sait; Gürdal, Mehmet; Palamar, Melis; Çavuşoğlu, Türker; Ateş, Utku; Veral, Ali; Güven, Çağrı; Wolosin, Jose Mario

2017-10-01

The cornea is the outermost tissue of the eye and it must be transparent for the maintenance of good visual function. The superficial epithelium of the cornea, which is renewed continuously by corneal stem cells, plays a critical role in the permanence of this transparency. These stem cells are localized at the cornea-conjunctival transition zone, referred to as the limbus. When this zone is affected/destroyed, limbal stem cell deficiency ensues. Loss of limbal stem cell function allows colonization of the corneal surface by conjunctival epithelium. Over 6 million people worldwide are affected by corneal blindness, and limbal stem cell deficiency is one of the main causes. Fortunately, it is becoming possible to recover vision by autologous transplantation of limbal cells obtained from the contralateral eye in unilateral cases. Due to the potential risks to the donor eye, only a small amount of tissue can be obtained, in which only 1-2% of the limbal epithelial cells are actually limbal stem cells. Vigorous attempts are being made to expand limbal stem cells in culture to preserve or even enrich the stem cell population. Ex vivo expanded limbal stem cell treatment in limbal stem cell deficiency was first reported in 1997. In the 20 years since, various protocols have been developed for the cultivation of limbal epithelial cells. It is still not clear which method promotes effective stem cell viability and this remains a subject of ongoing research. The most preferred technique for limbal cell culture is the explant culture model. In this approach, a small donor eye limbal biopsy is placed as an explant onto a biocompatible substrate (preferably human amniotic membrane) for expansion. The outgrowth (cultivated limbal epithelial cells) is then surgically transferred to the recipient eye. Due to changing regulations concerning cell-based therapy, the implementation of cultivated limbal epithelial transplantation in accordance with Good Laboratory Practice using xenobiotic-free systems is becoming widely accepted both in Turkey and worldwide.

Optimized human platelet lysate as novel basis for a serum-, xeno-, and additive-free corneal endothelial cell and tissue culture.

PubMed

Thieme, Daniel; Reuland, Lynn; Lindl, Toni; Kruse, Friedrich; Fuchsluger, Thomas

2018-02-01

The expansion of donor-derived corneal endothelial cells (ECs) is a promising approach for regenerative therapies in corneal diseases. To achieve the best Good Manufacturing Practice standard the entire cultivation process should be devoid of nonhuman components. However, so far, there is no suitable xeno-free protocol for clinical applications. We therefore introduce a processed variant of a platelet lysate for the use in corneal cell and tissue culture based on a Good Manufacturing Practice-grade thrombocyte concentrate. This processed human platelet lysate (phPL), free of any animal components and of anticoagulants such as heparin with a physiological ionic composition, was used to cultivate corneal ECs in vitro and ex vivo in comparison to standard cultivation with fetal calf serum (FCS). Human donor corneas were cut in quarters while 2 quarters of each cornea were incubated with the respective medium supplement. Three fields of view per quarter were taken into account for the analysis. Evaluation of phPL as a medium supplement in cell culture of immortalized EC showed a superior viability compared with FCS control with reduced cell proliferation. Furthermore, the viability during the expansion of primary cells is significantly (3-fold ±0.5) increased with phPL compared with FCS standard medium. Quartering donor corneas was traumatic for the endothelium and therefore resulted in increased EC loss. Interestingly, however, cultivation of the quartered pieces for 2 weeks in 0.1-mg/ml pHPL in Biochrome I showed a 21 (±10) % EC loss compared with 67 (±12) % EC loss when cultivated in 2% FCS in Biochrome I. The cell culture protocol with pHPL as FCS replacement seems to be superior to the standard FCS protocols with respect to EC survival. It offers a xeno-free and physiological environment for corneal endothelial cells. This alternative cultivation protocol could facilitate the use of EC for human corneal cell therapy. Copyright © 2017 John Wiley & Sons, Ltd.

Quercetin and the ocular surface: What we know and where we are going

PubMed Central

McKay, Tina B

2017-01-01

Flavonoids are a class of plant and fungus secondary metabolites that serve functional roles in protecting against UV-induced oxidative stress, mediating auxin signaling, and promoting microbial defense. Flavonoids are extremely abundant in nature where their potent antioxidant capacity and very low toxicity makes them highly attractive as potential therapeutic agents. In terms of clinical applications, neither the Food and Drug Administration (FDA) nor the European Food Safety Authority (EFSA) has approved any health claims or drugs related to the use of flavonoids for therapeutic purposes. Quercetin is a common flavonol that has been shown to have potent antioxidant, anti-inflammatory, and anti-fibrotic activities both in vitro and in vivo in various tissues. Recently, the application of quercetin as a therapeutic has been gaining attention in the ocular surface scientific community in the study of dry eye, keratoconus, inflammation, and neovascularization of the cornea. This review will discuss the latest findings and the use of quercetin for the treatment of dystrophies of the ocular surface. Impact statement The eye represents a small portion of the human body, accounting for one decimal fraction of the anterior body surface. The cornea is an avascular, transparent tissue that acts as a primary barrier against mechanical and infectious damaging agents, protecting the internal structures of the eye. Corneal survival and function are affected by a number of factors including but not limited to injury, trauma, infection, genetics, and environment. Corneal injury, or trauma, often leads to loss of corneal transparency and even blindness. The concept of “curing” corneal opacity has been discussed in published form for over 200 years. Currently, full corneal transplant is the only treatment option. There is a strong interest in developing natural therapeutic products that come with minimum side effects. A novel antioxidant flavonoid, quercetin, has been gaining traction as a potential therapeutic to prevent the injured cornea. This review discusses the potential of this antioxidant. PMID:28056553

Quercetin and the ocular surface: What we know and where we are going.

PubMed

McKay, Tina B; Karamichos, Dimitrios

2017-03-01

Flavonoids are a class of plant and fungus secondary metabolites that serve functional roles in protecting against UV-induced oxidative stress, mediating auxin signaling, and promoting microbial defense. Flavonoids are extremely abundant in nature where their potent antioxidant capacity and very low toxicity makes them highly attractive as potential therapeutic agents. In terms of clinical applications, neither the Food and Drug Administration (FDA) nor the European Food Safety Authority (EFSA) has approved any health claims or drugs related to the use of flavonoids for therapeutic purposes. Quercetin is a common flavonol that has been shown to have potent antioxidant, anti-inflammatory, and anti-fibrotic activities both in vitro and in vivo in various tissues. Recently, the application of quercetin as a therapeutic has been gaining attention in the ocular surface scientific community in the study of dry eye, keratoconus, inflammation, and neovascularization of the cornea. This review will discuss the latest findings and the use of quercetin for the treatment of dystrophies of the ocular surface. Impact statement The eye represents a small portion of the human body, accounting for one decimal fraction of the anterior body surface. The cornea is an avascular, transparent tissue that acts as a primary barrier against mechanical and infectious damaging agents, protecting the internal structures of the eye. Corneal survival and function are affected by a number of factors including but not limited to injury, trauma, infection, genetics, and environment. Corneal injury, or trauma, often leads to loss of corneal transparency and even blindness. The concept of "curing" corneal opacity has been discussed in published form for over 200 years. Currently, full corneal transplant is the only treatment option. There is a strong interest in developing natural therapeutic products that come with minimum side effects. A novel antioxidant flavonoid, quercetin, has been gaining traction as a potential therapeutic to prevent the injured cornea. This review discusses the potential of this antioxidant.

Geometrical gradients in the distribution of temperature and absorbed ultraviolet radiation in ocular tissues.

PubMed

Sliney, David H

2002-01-01

The geographical variations in the incidence of age-related ocular changes such as presbyopia and cataracts and diseases such as pterygium and droplet keratopathies have led to theories pointing to sunlight, ultraviolet radiation (UVR) exposure and ambient temperature as potential etiological factors. Some epidemiological evidence also points to an association of age-related macular degeneration to sunlight exposure. The actual distribution of sunlight exposure and the determination of temperature variations of different tissues within the anterior segment of the eye are difficult to assess. Of greatest importance are the geometrical factors that influence selective UVR exposures to different segments of the lens, cornea and retina. Studies show that the temperature of the lens and cornea varies by several degrees depending upon climate, and that the incidence of nuclear cataract incidence is greater in areas of higher ambient temperature (i.e., in the tropics). Likewise, sunlight exposure to local areas of the cornea, lens and retina varies greatly in different environments. However, epidemiological studies of the influence of environmental UVR in the development of cataract, pterygium, droplet keratopathies and age-related macular degeneration have produced surprisingly inconsistent findings. The lack of consistent results is seen to be due largely to either incomplete or erroneous estimates of outdoor UV exposure dose. Geometrical factors dominate the determination of UVR exposure of the eye. The degree of lid opening limits ocular exposure to rays entering at angles near the horizon. Clouds redistribute overhead UVR to the horizon sky. Mountains, trees and building shield the eye from direct sky exposure. Most ground surfaces reflect little UVR. The result is that highest UVR exposure occurs during light overcast where the horizon is visible and ground surface reflection is high. By contrast, exposure in a high mountain valley (lower ambient temperature) with green foliage results in a much lower ocular dose. Other findings of these studies show that retinal exposure to light and UVR in daylight occurs largely in the superior retina.

Cryopreservation of Living Organs

NASA Astrophysics Data System (ADS)

Tanasawa, Ichiro; Nagata, Shinichi; Kimura, Naohiro

Cryopreservation is considered to be the most promising way of preserving living organs or tissues for a long period of time without casuing any damage to their biological functions. However, cryopreservation has been succeeded only for simple and small-size tissues such as spermatozoon, ovum, erythrocyte, bone marrow and cornea. Cryopreservation of more complex and large-scale organs are not yet succssful. The authors have attempted to establish a technique for cryopreservation of larger living organs. An experiment was carried out using daphnia (water flea). The optimum rates of freezing and thawing were determined together with the optimum selection of cryoprotectant. High recovery rate was achieved under these conditions.

Obtaining corneal tissue for keratoplasty.

PubMed

Navarro Martínez-Cantullera, A; Calatayud Pinuaga, M

2016-10-01

Cornea transplant is the most common tissue transplant in the world. In Spain, tissue donation activities depend upon transplant coordinator activities and the well-known Spanish model for organ and tissue donation. Tissue donor detection system and tissue donor evaluation is performed mainly by transplant coordinators using the Spanish model on donation. The evaluation of a potential tissue donor from detection until recovery is based on an exhaustive review of the medical and social history, physical examination, family interview to determine will of the deceased, and a laboratory screening test. Corneal acceptance criteria for transplantation have a wider spectrum than other tissues, as donors with active malignancies and infections are accepted for kearatoplasty in most tissue banks. Corneal evaluation during the whole process is performed to ensure the safety of the donor and the recipient, as well as an effective transplant. Last step before processing, corneal recovery, must be performed under standard operating procedures and in a correct environment. Copyright © 2016 Sociedad Española de Oftalmología. Published by Elsevier España, S.L.U. All rights reserved.

Transient downregulation of microRNA-206 protects alkali burn injury in mouse cornea by regulating connexin 43

PubMed Central

Li, Xiaoyan; Zhou, Huanfen; Tang, Weiqiang; Guo, Qing; Zhang, Yan

2015-01-01

Purpose: Chemical burn in cornea may cause permanent visual problem or complete blindness. In the present study, we investigated the role of microRNA 206 (miR-206) in relieving chemical burn in mouse cornea. Method: An alkali burn model was established in C57BL/6 mice to induce chemical corneal injury. Within 72 hours, the transient inflammatory responses in alkali-treated corneas were measured by opacity and corneal neovascularization (CNV) levels, and the gene expression profile of miR-206 was measured by quantitative real-time PCR (qPCR). Inhibitory oligonucleotides of miR-206, miR-206-I, were intrastromally injected into alkali-burned corneas. The possible protective effects of down-regulating miR-206 were assessed by both in vivo measurements of inflammatory responses and in vitro histochemical examinations of corneal epithelium sections. The possible binding of miR-206 on its molecular target, connexin43 (Cx43), was assessed by luciferase reporter (LR) and western blot (WB) assays. Cx43 was silenced by siRNA to examine its effect on regulating miR-206 modulation in alkali-burned cornea. Results: Opacity and CNV levels, along with gene expression of miR-206, were all transiently elevated within 72 hours of alkali-burned mouse cornea. Intrastromal injection of miR-206-I into alkali-burned cornea down-regulated miR-206 and ameliorated inflammatory responses both in vivo and in vitro. LR and WB assays confirmed that Cx43 was directly targeted by miR-206 in mouse cornea. Genetic silencing of Cx43 reversed the protective effect of miR-206 down-regulation in alkali-burned cornea. Conclusion: miR-206, associated with Cx43, is a novel molecular modulator in alkali burn in mouse cornea. PMID:26045777

The use of human cornea organotypic cultures to study herpes simplex virus type 1 (HSV-1)-induced inflammation.

PubMed

Drevets, Peter; Chucair-Elliott, Ana; Shrestha, Priyadarsini; Jinkins, Jeremy; Karamichos, Dimitrios; Carr, Daniel J J

2015-10-01

To determine the utility of human organotypic cornea cultures as a model to study herpes simplex virus type 1 (HSV-1)-induced inflammation and neovascularization. Human organotypic cornea cultures were established from corneas with an intact limbus that were retrieved from donated whole globes. One cornea culture was infected with HSV-1 (10(4) plaque-forming units), while the other cornea from the same donor was mock-infected. Supernatants were collected at intervals post-culture with and without infection to determine viral titer (by plaque assay) and pro-angiogenic and proinflammatory cytokine concentration by suspension array analysis. In some experiments, the cultured corneas were collected and evaluated for HSV-1 antigens by immunohistochemical means. Another set of experiments measured susceptibility of human three-dimensional cornea fibroblast constructs, in the presence and absence of TGF-β1, to HSV-1 infection in terms of viral replication and the inflammatory response to infection as a comparison to the organotypic cornea cultures. Organotypic cornea cultures and three-dimensional fibroblast constructs exhibited varying degrees of susceptibility to HSV-1. Fibroblast constructs were more susceptible to infection in terms of infectious virus recovered in a shorter period of time. There were changes in the levels of select pro-angiogenic or proinflammatory cytokines that were dictated as much by the cultures producing them as by whether they were infected with HSV-1 or treated with TGF-β1. Organotypic cornea and three-dimensional fibroblast cultures are likely useful for the identification and short-term study of novel antiviral compounds and virus replication, but are limited in the study of the local immune response to infection.

Eye Irritation Test (EIT) for Hazard Identification of Eye Irritating Chemicals using Reconstructed Human Cornea-like Epithelial (RhCE) Tissue Model.

PubMed

Kaluzhny, Yulia; Kandárová, Helena; d'Argembeau-Thornton, Laurence; Kearney, Paul; Klausner, Mitchell

2015-08-23

To comply with the Seventh Amendment to the EU Cosmetics Directive and EU REACH legislation, validated non-animal alternative methods for reliable and accurate assessment of ocular toxicity in man are needed. To address this need, we have developed an eye irritation test (EIT) which utilizes a three dimensional reconstructed human cornea-like epithelial (RhCE) tissue model that is based on normal human cells. The EIT is able to separate ocular irritants and corrosives (GHS Categories 1 and 2 combined) and those that do not require labeling (GHS No Category). The test utilizes two separate protocols, one designed for liquid chemicals and a second, similar protocol for solid test articles. The EIT prediction model uses a single exposure period (30 min for liquids, 6 hr for solids) and a single tissue viability cut-off (60.0% as determined by the MTT assay). Based on the results for 83 chemicals (44 liquids and 39 solids) EIT achieved 95.5/68.2/ and 81.8% sensitivity/specificity and accuracy (SS&A) for liquids, 100.0/68.4/ and 84.6% SS&A for solids, and 97.6/68.3/ and 83.1% for overall SS&A. The EIT will contribute significantly to classifying the ocular irritation potential of a wide range of liquid and solid chemicals without the use of animals to meet regulatory testing requirements. The EpiOcular EIT method was implemented in 2015 into the OECD Test Guidelines as TG 492.

Eye Irritation Test (EIT) for Hazard Identification of Eye Irritating Chemicals using Reconstructed Human Cornea-like Epithelial (RhCE) Tissue Model

PubMed Central

Kaluzhny, Yulia; Kandárová, Helena; d’Argembeau-Thornton, Laurence; Kearney, Paul; Klausner, Mitchell

2015-01-01

To comply with the Seventh Amendment to the EU Cosmetics Directive and EU REACH legislation, validated non-animal alternative methods for reliable and accurate assessment of ocular toxicity in man are needed. To address this need, we have developed an eye irritation test (EIT) which utilizes a three dimensional reconstructed human cornea-like epithelial (RhCE) tissue model that is based on normal human cells. The EIT is able to separate ocular irritants and corrosives (GHS Categories 1 and 2 combined) and those that do not require labeling (GHS No Category). The test utilizes two separate protocols, one designed for liquid chemicals and a second, similar protocol for solid test articles. The EIT prediction model uses a single exposure period (30 min for liquids, 6 hr for solids) and a single tissue viability cut-off (60.0% as determined by the MTT assay). Based on the results for 83 chemicals (44 liquids and 39 solids) EIT achieved 95.5/68.2/ and 81.8% sensitivity/specificity and accuracy (SS&A) for liquids, 100.0/68.4/ and 84.6% SS&A for solids, and 97.6/68.3/ and 83.1% for overall SS&A. The EIT will contribute significantly to classifying the ocular irritation potential of a wide range of liquid and solid chemicals without the use of animals to meet regulatory testing requirements. The EpiOcular EIT method was implemented in 2015 into the OECD Test Guidelines as TG 492. PMID:26325674

Ocular hazards of the colors used during the festival-of-colors (Holi) in India--malachite green toxicity.

PubMed

Velpandian, T; Saha, K; Ravi, A K; Kumari, S S; Biswas, N R; Ghose, S

2007-01-10

The objective of this study was to evaluate the nature of the colors used and their toxicity to the eye upon exposure to them during celebration of Holi (our festival-of-colors). Color powders and formulations were procured at random in and around Delhi during the festival. The green/bluish-green colors reported with the higher incidence of ocular toxicity were subjected for further evaluation. Eyewash fluid collected from the patients exposed to the colors was also subjected for analysis. This study was further extended to evaluate the corneal penetration of malachite green using goat cornea in perfusion chamber. In 16/18 color samples collected, malachite green or 4-[(4-dimethylaminophenyl)-phenyl-methyl]-N,N-dimethyl-aniline was detected at different concentrations. In the eyewash fluid of four patients, HPLC estimation confirmed the presence of malachite green at concentrations of 1.3, 0.18, 3.5 and 5.4 microg in 250 ml which was responsible for its reported toxicity. The in vitrotrans-corneal penetration studies did not show any detectable amount of malachite green in effluent fluid-in vitro tissue retention studies revealed that increasing the contact time increases tissue concentration. After 2 min of exposure, the tissue concentration was significantly higher. To conclude, malachite green was extensively used in our festival of Holi and has caused severe ocular irritation with epithelial defect upon exposure, though it did not penetrate through the cornea-further in vitro and in vivo studies are required on colors used in Holi.

Ocular surface changes in limbal stem cell deficiency caused by chemical injury: a histologic study of excised pannus from recipients of cultured corneal epithelium.

PubMed

Fatima, A; Iftekhar, G; Sangwan, V S; Vemuganti, G K

2008-09-01

To report histopathologic changes of the ocular surface pannus in patients with severe limbal stem cell deficiency (LSCD). Corneal and conjunctival pannus tissues from 29 patients undergoing ocular reconstruction with cultured limbal cell transplantation were included. The medical records of these patients were reviewed for demographics, aetiologic diagnosis, type of injury, interval between the initial insult and excision of pannus, and medical history involving human amniotic membrane (HAM) or limbal transplantation. The paraffin-embedded tissues were reviewed for epithelial changes, type-degree of fibrosis, degenerative changes, vascular changes, conjunctivalization of corneal surface, and evidence of residual HAM. We attempted a clinicopathologic correlation to understand the pathogenesis of pannus formation in LSCD. The 29 tissues were from 29 eyes of patients with primary aetiology of chemical burn in 89.6% (undetermined in 10.4%) of cases. The pannus showed epithelial hyperplasia in 62%, active fibrosis in 66%, severe inflammation in 21%, giant cell reaction in 28%, and stromal calcification in 14% cases. Goblet cells were seen over the cornea in 64% cases; their absence was associated with squamous metaplasia of the conjunctiva and with long duration of insult. Evidence of residual HAM was noted in 42% cases. The commonest cause of severe LSCD is alkali-induced injury. Goblet cells over the cornea were seen in 60% of cases. HAM used for ocular surface reconstruction could persist for long periods within the corneal pannus, thus raising the need for further studies with long-term follow-up.

Effect of intracameral injection of fibrin tissue sealant on the rabbit anterior segment

PubMed Central

Chew, Annabel C.Y.; Tan, Donald T.H.; Poh, Rebekah; HM, Htoon; Beuerman, Roger W.

2010-01-01

Purpose To investigate the effect of intracameral injection of fibrin tissue sealant on the anterior segment structures in a rabbit model. Methods One eye of 10 rabbits received an intracameral injection of fibrin tissue sealant with a thrombin concentration of 500 IU (TISSEEL), and the fellow eye received an intracameral injection of balanced salt solution as a control. The rabbits were followed up with serial slit-lamp examinations, photography, high resolution anterior segment optical coherence tomography scans with pachymetry measurement, and intraocular pressure (IOP) monitoring until complete dissolution of the fibrin sealant. Corneal endothelial cell viability was evaluated using live/dead cell assays. Apoptosis of the cornea and trabecular meshwork were evaluated using TUNEL assays. Ultra-structural examinations of the cornea and trabecular meshwork were performed using electron microscopy. Histology of the trabecular meshwork and iris were analyzed using light microscopy. Results The quantity of the intracameral fibrin sealant was shown to be significantly correlated with increased IOP and pachymetry post-operatively. Complete dissolution of the fibrin sealant occurred between 15 and 30 days. Live/dead cell assays showed no decrease in viability of the corneal endothelium, and TUNEL assays showed no increase in apoptosis of the corneal epithelium, stroma, endothelium, or trabecular meshwork in the eyes with the fibrin sealant. Light and electron microscopy of the anterior segment structures were unremarkable. Conclusion The intracameral use of fibrin glue was associated with a transient increase in IOP and pachymetry. However, there was no evidence of toxicity or structural damage to the corneal endothelium, trabecular meshwork, or iris. PMID:20596250

Human fibrin tissue adhesive for sutureless lamellar keratoplasty and scleral patch adhesion: a pilot study.

PubMed

Kaufman, Herbert E; Insler, Michael S; Ibrahim-Elzembely, Hosan A; Kaufman, Stephen C

2003-11-01

To determine whether a fibrin adhesive can facilitate the performance of sutureless lamellar keratoplasty and attachment of amnion to bare sclera. Prospective, noncomparative case series. Six patients, 5 of whom underwent lamellar keratoplasty and 1 who received an amniotic patch of the sclera and cornea. Institutional review board approval was not required for these therapeutic treatments. In 5 patients, the epithelium was removed from the corneal surface, a free cap, 200- micro m thick, was cut with a microkeratome, and a human fibrin tissue adhesive (Tisseel VH Fibrin Sealant; Baxter Healthcare Corporation, Glendale, CA) was applied to the cut surface of the corneal stroma. A 200- micro m thick, microkeratome-cut lamellar graft was placed in the stromal bed without sutures, and a bandage soft contact lens was applied. The lens was left in place for 1 week and then removed. In 1 patient, the adhesive was applied to bare sclera for attachment of amniotic membrane after removal of a conjunctival melanosis. All patients were followed up for 3 months after surgery. Tissue adhesion, corneal clarity, and visual acuity. All 5 lamellar grafts healed and remained clear, although final visual acuity varied with visual potential and astigmatism. The amniotic membrane graft also adhered well to the bare sclera. The fibrin adhesive provided satisfactory attachment without sutures for lamellar keratoplasty and amniotic patching. It should be effective for sealing of clear cornea incisions, LASIK flaps, and conjunctival and skin grafts. An adhesive that has been designed specifically for ophthalmic applications and is more convenient to use would be desirable.

Investigations of the corneal epithelium in Veterinary Medicine: State of the art on corneal stem cells found in different mammalian species and their putative application.

PubMed

Patruno, M; Perazzi, A; Martinello, T; Gomiero, C; Maccatrozzo, L; Iacopetti, I

2018-05-08

The existence of progenitor cells that can readily differentiate into a specific cell type is a common cellular strategy for physiological tissue growth and repair mechanisms. In the mammalian cornea, many aspects regarding the nature and location of these cells are still unclear. In the human limbus (peripheral area of the cornea) progenitor cells have been found and characterized but in non-human mammals, the picture is not so clear. In this review, we examine current knowledge about the morphology of limbus and the localization of corneal epithelial stem cells in all species studied so far, comparing data with humans. We have also explored different research directions in the veterinary field in order to discuss the: i) currently used protocols and ii) best range of treatments for ocular pathologies in which corneal stem cells are involved. Copyright © 2018. Published by Elsevier Ltd.

Multiphoton tomography of the human eye

NASA Astrophysics Data System (ADS)

König, Karsten; Batista, Ana; Hager, Tobias; Seitz, Berthold

2017-02-01

Multiphoton tomography (MPT) is a novel label-free clinical imaging method for non-invasive tissue imaging with high spatial (300 nm) and temporal (100 ps) resolutions. In vivo optical histology can be realized due to the nonlinear excitation of endogenous fluorophores and second-harmonic generation (SHG) of collagen. Furthermore, optical metabolic imaging (OMI) is performed by two-photon autofluorescence lifetime imaging (FLIM). So far, applications of the multiphoton tomographs DermaInspect and MPTflex were limited to dermatology. Novel applications include intraoperative brain tumor imaging as well as cornea imaging. In this work we describe two-photon imaging of ex vivo human corneas unsuitable for transplantation. Furthermore, the cross-linking (CXL) process of corneal collagen based on UVA exposure and 0.1 % riboflavin was studied. The pharmacokinetics of the photosensitizer could be detected with high spatial resolution. Interestingly, an increase in the stromal autofluorescence intensity and modifications of the autofluorescence lifetimes were observed in the human corneal samples within a few days following CXL.

Limbal explants from cryopreserved cadaver human corneas. Immunofluorescence and light microscopy of epithelial cells growing in culture.

PubMed

Bratanov, M; Neronov, A; Nikolova, E

2009-01-01

The aim of the present study was to determine whether human cadaver corneas, that were subject to cryopreservation, would be a source of migrating epithelial cells in vitro and what kind of morphological features these cells possess. Limbal explant culture was used for expanding the epithelial cells. Non-quantitative light microscopical examinations of the cultures within a period of 28 days were carried out. The phenotype of cultured cells, particularly of the presumed adult stem cell population, was examined by indirect fluorescent immunostaining using antibodies against corneal stem cell associated markers p63 and vimentin. The effectiveness of the freezing-thawing protocol was confirmed by cultivation of limbal explants taken from non-cryopreserved cadaver corneoscleral rims. The result clearly showed that limbal tissue, subjected to cryopreservation and long lasting (up to 12 months) storage in liquid nitrogen, retains the capacity to be source of migrating and proliferating epithelial cells in vitro including the presumed adult stem cells and transient amplifying cells.

Determination of excimer laser ablation rate of the human cornea using in vivo Scheimpflug videography.

PubMed

Huebscher, H J; Genth, U; Seiler, T

1996-01-01

To determine in vivo the amount of human corneal tissue removed by each excimer laser pulse, the so-called ablation rate, during photorefractive keratectomy (PRK). There is confusion in the literature because the experimentally determined ablation rate of 0.4 to 0.5 microns per pulse differs from the nominal ablation rate of 0.23 to 0.3 microns per pulse, which is the value used in clinical procedures. Eleven eyes of 11 patients were treated with PRK for correction of myopia. The corneal curvature was determined by Scheimpflug videography before and immediately after surgery. Starting from this curvature change, the authors calculated the real ablation rate. The real ablation rate is coincident with the nominal ablation rate and differs significantly from the ablation rate derived from deep keratectomy experiments. The outer layers of the cornea show significantly different ablation behavior than the deeper stroma. This information has clinical relevance for the predictability of intrastromal excimer laser procedures.

Deep lamellar keratoplasty on air with lyophilised tissue.

PubMed Central

Chau, G K; Dilly, S A; Sheard, C E; Rostron, C K

1992-01-01

Deep lamellar keratoplasty on air involves injecting air into the corneal stroma to expand it to several times its normal thickness. This method is designed to facilitate dissection of the deep stroma and reduce the risk of perforation of Descemet's membrane when carrying out deep lamellar keratoplasty. We have modified the technique by using prelathed freeze-dried donor tissue and report our results in a series of patients with corneal stromal scarring owing to a variety of corneal problems, namely, keratoconus, pterygium, and herpes zoster ophthalmicus. All patients achieved best corrected postoperative visual acuity of 6/12 or better without problems associated with graft failure or rejection. Histopathological examination of corneal tissue following air injection showed surgical emphysema within the cornea and separation of deep stromal fibres from the underlying Descemet's membrane. Images PMID:1477037

Protective effect of 4-coumaric acid from UVB ray damage in the rabbit eye.

PubMed

Lodovici, Maura; Caldini, Silvia; Morbidelli, Lucia; Akpan, Victor; Ziche, Marina; Dolara, Piero

2009-01-08

UV-induced oxidation damage seems to play a major role in a number of specific pathological conditions of intraocular tissues, such as cataract formation and retinal degeneration. Therefore, antioxidant and/or scavenger compounds might protect the eyes from UV-induced cellular damage. We previously reported that 4-coumaric acid (4-CA) is able to protect rabbit corneal-derived cells (SIRC) from UVB-induced oxidation damage. In this study we evaluated the protective effect of 4-CA against UVB-induced cell damage in rabbit cornea in vivo. Twelve male New Zealand albino rabbits were used; four rabbits were used as a control and received vehicle in one eye and 4-CA acid in the contralateral eye; eight rabbits were exposed to UVB rays (79.2mJ/cm(2)) and three days before to UV exposure each animal received 1 drop/day of vehicle in one eye and 1 drop/day of vehicle containing 4-CA (164ng) in the contralateral eye. Corneal and sclera tissues were removed and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) levels were measured. Superoxide dismutase (SOD) and xanthine oxidase (XO) activities were determined in aqueous humour. UVB-induced vessel hyper-reactivity was strongly reduced at 4 and 24h after UVB exposure after local treatment with 4-CA, 8-oxodGuo levels, a marker of oxidative DNA damage, were significantly increased (P<0.05) in sclera and cornea by UVB irradiation, but when 4-CA was administered to the conjunctiva in a buffered solution once a day for 3d before and 6d after UVB exposure, levels of 8-oxodGuo were similar to controls and significantly reduced (P<0.05) compared to UVB-treated corneas. XO activity in the aqueous humour was significantly increased. The administration of 4-CA for 3d before and 6d after UVB irradiation induced a small but significant (P<0.05) reduction of XO compared with control eyes. Our results indicate that the administration of 4-CA protects eye tissues, thus reducing the harmful effect of UVB radiation at low concentration, probably through its free radical scavenging and antioxidant properties. Therefore, 4-CA may be useful in protecting the eye from free radical damage following UVB exposure from sunlight, UV lamps and welding torches.

Evaluating the Effects of Riboflavin/UV-A and Rose-Bengal/Green Light Cross-Linking of the Rabbit Cornea by Noncontact Optical Coherence Elastography

PubMed Central

Singh, Manmohan; Li, Jiasong; Han, Zhaolong; Vantipalli, Srilatha; Liu, Chih-Hao; Wu, Chen; Raghunathan, Raksha; Aglyamov, Salavat R.; Twa, Michael D.; Larin, Kirill V.

2016-01-01

Purpose The purpose of this study was to use noncontact optical coherence elastography (OCE) to evaluate and compare changes in biomechanical properties that occurred in rabbit cornea in situ after corneal collagen cross-linking by either of two techniques: ultraviolet-A (UV-A)/riboflavin or rose-Bengal/green light. Methods Low-amplitude (≤10 μm) elastic waves were induced in mature rabbit corneas by a focused air pulse. Elastic wave propagation was imaged by a phase-stabilized swept source OCE (PhS-SSOCE) system. Corneas were then cross-linked by either of two methods: UV-A/riboflavin (UV-CXL) or rose-Bengal/green light (RGX). Phase velocities of the elastic waves were fitted to a previously developed modified Rayleigh-Lamb frequency equation to obtain the viscoelasticity of the corneas before and after the cross-linking treatments. Micro-scale depth-resolved phase velocity distribution revealed the depth-wise heterogeneity of both cross-linking techniques. Results Under standard treatment settings, UV-CXL significantly increased the stiffness of the corneas by ∼47% (P < 0.05), but RGX did not produce statistically significant increases. The shear viscosities were unaffected by either cross-linking technique. The depth-wise phase velocities showed that UV-CXL affected the anterior ∼34% of the corneas, whereas RGX affected only the anterior ∼16% of the corneas. Conclusions UV-CXL significantly strengthens the cornea, whereas RGX does not, and the effects of cross-linking by UV-CXL reach deeper into the cornea than cross-linking effects of RGX under similar conditions. PMID:27409461

Screening for previous refractive surgery in eye bank corneas by using optical coherence tomography.

PubMed

Lin, Roger C; Li, Yan; Tang, Maolong; McLain, Marcy; Rollins, Andrew M; Izatt, Joseph A; Huang, David

2007-06-01

To use optical coherence tomography (OCT) to detect previous refractive surgery in donor corneas. We constructed a tabletop OCT scanner operating at 1310-nm wavelength. Donor corneas at the Cleveland Eye Bank were scanned while sealed within the sterile container immersed in Optisol GS. OCT scanning was performed with 7.6-mm-long lines (512 axial scans) along 8 meridians. Anterior and posterior corneal surfaces were automatically mapped using image processing software that we developed. Curvature was computed from the best parabolic fit in the central 5-mm diameter. Layered analysis of the stromal reflectivity was also performed. Twenty-nine corneas from 19 donors were examined. Five had a history of laser in situ keratomileusis (LASIK). The flap interfaces could not be visualized on slit-lamp or OCT images but were confirmed by histology. The death-to-scan time was 22.1 +/- 11.4 (SD) hours for normal corneas and 100.6 +/- 57.5 hours for LASIK corneas. The anterior surface power was 67.5 +/- 2.5 D in control corneas and 64.5 +/- 2.4 D in LASIK corneas (P = 0.023). There was no significance between the 2 groups in terms of posterior curvature and thickness parameters. The anterior/posterior reflectivity ratio in the central 4-mm diameter was significantly lower in post-LASIK corneas than in control (P < 0.05). OCT provides thickness, topography, and reflectivity maps of donor corneas without taking them out of preservation medium and container. The anterior curvature and the anterior/posterior stromal reflectivity ratio may be useful for detecting previous LASIK.

Femtosecond lasers in ophthalmology: clinical applications in anterior segment surgery

NASA Astrophysics Data System (ADS)

Juhasz, Tibor; Nagy, Zoltan; Sarayba, Melvin; Kurtz, Ronald M.

2010-02-01

The human eye is a favored target for laser surgery due to its accessibility via the optically transparent ocular tissue. Femtosecond lasers with confined tissue effects and minimized collateral tissue damage are primary candidates for high precision intraocular surgery. The advent of compact diode-pumped femtosecond lasers, coupled with computer controlled beam delivery devices, enabled the development of high precision femtosecond laser for ophthalmic surgery. In this article, anterior segment femtosecond laser applications currently in clinical practice and investigation are reviewed. Corneal procedures evolved first and remain dominant due to easy targeting referenced from a contact surface, such as applanation lenses placed on the eye. Adding a high precision imaging technique, such as optical coherence tomography (OCT), can enable accurate targeting of tissue beyond the cornea, such as the crystalline lens. Initial clinical results of femtosecond laser cataract surgery are discussed in detail in the latter portion part of the article.

Thermal damage produced by high-irradiance continuous wave CO2 laser cutting of tissue.

PubMed

Schomacker, K T; Walsh, J T; Flotte, T J; Deutsch, T F

1990-01-01

Thermal damage produced by continuous wave (cw) CO2 laser ablation of tissue in vitro was measured for irradiances ranging from 360 W/cm2 to 740 kW/cm2 in order to investigate the extent to which ablative cooling can limit tissue damage. Damage zones thinner than 100 microns were readily produced using single pulses to cut guinea pig skin as well as bovine cornea, aorta, and myocardium. Multiple pulses can lead to increased damage. However, a systematic decrease in damage with irradiance, predicted theoretically by an evaporation model of ablation, was not observed. The damage-zone thickness was approximately constant around the periphery of the cut, consistent with the existence of a liquid layer which stores heat and leads to tissue damage, and with a model of damage and ablation recently proposed by Zweig et al.

[Biomechanical condition of the cornea as a new indicator for pathological and structural changes].

PubMed

Spörl, E; Terai, N; Haustein, M; Böhm, A G; Raiskup-Wolf, F; Pillunat, L E

2009-06-01

Several methods permit the measurement of geometric parameters of the cornea, but until now biomechanical conditions of the cornea have been ignored (e.g. in refractive corneal surgery). Besides the geometric condition, biomechanical properties of the cornea have been shown to influence applanation measurement of intra-ocular pressure (IOP) and epidemiological studies have identified corneal thickness as an independent risk factor for the development and progression of glaucoma. The aim of this investigation was to characterize the biomechanical properties of the cornea using the ocular response analyzer (ORA). The ocular response analyzer (ORA) is a new method available for non-contact measurement of the biomechanical properties of the cornea. We evaluated the reproducibility of measurements, the difference between static and dynamic factors and the impact of independent factors (e.g. IOP, age, CCT, swelling of the cornea) on 2,500 measurements of corneal hysteresis (CH) and corneal resistance factor (CRF). In a large sample size we observed changes in CH and CRF after refractive surgery procedures (LASIK, UV-A cross-linking, keratoplasty) and in other corneal disorders (keratoconus, corneal dystrophies). CRF and CH changes may reflect structural changes of the cornea. Thus, the ORA provides valuable information for a better understanding and characterization of the biomechanical condition of the cornea, especially with regard to diseases such as keratoconus and glaucoma.

Characterization of Rabbit Corneas Subjected to Stromal Stiffening by the Açaí Extract (Euterpe oleracea).

PubMed

Bersanetti, Patrícia A; Bueno, Tatiane L N; Morandim-Giannetti, Andreia de A; Nogueira, Regina F; Matos, Jivaldo R; Schor, Paulo

2017-04-01

In this study, we characterized rabbit corneas subjected to corneal cross-linking (CXL) with açaí extract compared with a riboflavin photo-stimulated procedure. The corneas of the slaughterhouse rabbits were divided into three groups: control, consisting of untreated corneal samples; riboflavin/UVA, where corneas were treated with 0.1% riboflavin photo-stimulated at 365 nm as the standard protocol; and açaí, where the samples were subjected to 4% açaí extract for 0.5-2 h. After the CXL procedure, corneas of the three groups were characterized by analyzing their elastic modulus and thermal denaturation profile. The elastic modulus at 3% strain showed an approximately threefold increase in the riboflavin/UVA group and 10.5 times in the corneas treated with 4% açaí extract for 2 h, compared with the control group (p < 0.01). The denaturation temperature values of the two groups of crosslinked corneas increased significantly (p < 0.05) and were more pronounced in the açaí group. The açaí extract was effective in promoting CXL in rabbit corneas as characterized by the different techniques.

In vivo confocal microscopy of human cornea covered with human amniotic membrane.

PubMed

Mimura, Tatsuya; Yamagami, Satoru; Usui, Tomohiko; Honda, Norihiko; Araki, Fumiyuki; Amano, Shiro

2008-01-01

Amniotic membrane transplantation has been widely performed to reconstruct the surface of the eye and treat chemical burns or epithelial defects. However, we have difficulty observing the cornea through the opaque transplanted amniotic membrane by slit-lamp biomicroscopy. We investigated the use of confocal microscopy for observation of human corneas covered with amniotic membrane. Human amniotic membrane was placed onto the normal corneas of five volunteers aged 22-24 years. Then, all layers of the covered corneas were observed by in vivo confocal microscopy. Confocal microscopy displayed the epithelium, basement membrane, and stroma of the amniotic membrane. It also displayed the corneal epithelium. Furthermore, corneal stromal keratocytes and the corneal endothelium were clearly observed through the amniotic membrane by confocal microscopy. We demonstrated that in vivo confocal microscopy enabled us to observe all layers of corneas covered with amniotic membrane in normal human eyes. Our findings suggest that confocal microscopy may have advantages for clinical examination of the ocular surface, including all layers of the cornea.

SPECIAL ISSUE DEVOTED TO MULTIPLE RADIATION SCATTERING IN RANDOM MEDIA: Optical coherent tomography measurements of the diffusion rate of water and drugs in an isolated and whole cornea

NASA Astrophysics Data System (ADS)

Larin, Kirill V.; Ghosn, M. G.

2006-12-01

The passive diffusion of drugs through the epithelial surfaces of an eye (the most widespread method for medical treatment of various diseases) is considered. The permeability of water and drugs through rabbit cornea was measured in the isolated cornea (separate from an eye) and in the whole cornea. The permeability coefficients of water and dexamethasone were estimated by the method of optical coherence tomography (OCT). Because multiple photon scattering introduces noise and distortions to the OCT signal, measurements were performed at depths up to 500 μm where most likely single scattering of light occurs in cornea. It is shown that the permeability coefficients in the isolated and whole cornea strongly differ from each other. For example, the water permeability in the isolated and whole cornea is (7.09±0.12)×10-5 and (1.71±0.51)×10-5 cm s-1, respectively.

Clinical and epidemiological aspects of cornea transplant patients of a reference hospital 1

PubMed Central

Cruz, Giovanna Karinny Pereira; de Azevedo, Isabelle Campos; Carvalho, Diana Paula de Souza Rego Pinto; Vitor, Allyne Fortes; Santos, Viviane Euzébia Pereira; Ferreira, Marcos Antonio

2017-01-01

ABSTRACT Objective: clinically characterizing cornea transplant patients and their distribution according to indicated and post-operative conditions of cornea transplantation, as well as estimating the average waiting time. Method: a cross-sectional, descriptive and analytical study performed for all cornea transplants performed at a reference service (n=258). Data were analyzed using Statistical Package for the Social Sciences, version 20.0. Results: the main indicator for cornea transplant was keratoconus. The mean waiting time for the transplant was approximately 5 months and 3 weeks for elective transplants and 9 days for urgent cases. An association between the type of corneal disorder with gender, age, previous surgery, eye classification, glaucoma and anterior graft failure were found. Conclusion: keratoconus was the main indicator for cornea transplant. Factors such as age, previous corneal graft failure (retransplantation), glaucoma, cases of surgeries prior to cornea transplant (especially cataract surgery) may be related to the onset corneal endothelium disorders. PMID:28614429

SkinEthic Laboratories, a company devoted to develop and produce in vitro alternative methods to animal use.

PubMed

de Brugerolle, Anne

2007-01-01

SkinEthic Laboratories is a France-based biotechnology company recognised as the world leader in tissue engineering. SkinEthic is devoted to develop and produce reliable and robust in vitro alternative methods to animal use in cosmetic, chemical and pharmaceutical industries. SkinEthic models provide relevant tools for efficacy and safety screening tests in order to support an integrated decision-making during research and development phases. Some screening tests are referenced and validated as alternatives to animal use (Episkin), others are in the process of validation under ECVAM and OECD guidelines. SkinEthic laboratories provide a unique and joined experience of more than 20 years from Episkin SNC and SkinEthic SA. Their unique cell culture process allows in vitro reconstructed human tissues with well characterized histology, functionality and ultrastructure features to be mass produced. Our product line includes skin models: a reconstructed human epidermis with a collagen layer, Episkin, reconstructed human epidermis without or with melanocytes (with a tanning degree from phototype II to VI) and a reconstructed human epithelium, i.e. cornea, and other mucosa, i.e. oral, gingival, oesophageal and vaginal. Our philosophy is based on 3 main commitments: to support our customers by providing robust and reliable models, to ensure training and education in using validated protocols, allowing a large array of raw materials, active ingredients and finished products in solid, liquid, powder, cream or gel form to be screened, and, to provide a dedicated service to our partners.

Stem cell roadmap - The industrial point of view.

PubMed

Elzaabi, Mazen; Thevenin, Agnès; Lirsac, Pierre-Noël

2017-01-01

CELLforCURE is a French Contract Development and Manufacturing Organization (CDMO) dedicated to industrialization and process development for routine manufacturing, GMP manufacturing for clinical and commercial batches and regulatory services and associated logistics. CELLforCURE is a subsidiary of LFB Group.Stem cells fields of application gather cell and gene therapy as well as tissue engineering. According to VisionGain survey, cell therapy medicinal products will remain predominant in the future.Clinical trials are sponsored either by universities or private companies. Most of clinical trials are performed in oncology (53%). More than 100 clinical trials are currently performed in France, involving 36 products in clinical phases II or II/III.Tomorrow's regenerative medicine will be organ reconstruction using scaffolds and bioprinting technologies. The expected applications in the near future could be skin, cornea, blood vessels, retina, urethra and trachea. There are still important issues to overcome: create the vasculature and neuron connection.Solutions are expected regarding I) fundamental biology, in particular better understanding of IPS behavior and metabolism, precursor differentiation conditions, sustainability of induced genetic changes, II) technical approaches which involves injectable preservation medium, high density cells and centrifugation system.

Corneal squamous cell carcinoma in a Border Collie.

PubMed

Busse, Claudia; Sansom, Jane; Dubielzig, R R; Hayes, Alison

2008-01-01

A 6-year-old, female, spayed Border Collie was presented to the Unit of Comparative Ophthalmology at the Animal Health Trust with a 6-month history of a progressive nonpainful opacity of the left cornea. A keratectomy was performed and the tissue submitted for histopathology. The diagnosis was squamous cell carcinoma. There has been no recurrence of the neoplasm to date (5 months). Canine corneal squamous cell carcinoma (SCC) has not been reported previously in the UK.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vogel, A.; Scammon, R.J.; Godwin, R.P.

Biological tissue is more susceptible to damage from tensile stress than to compressive stress. Tensile stress may arise through the thermoelastic response of laser-irradiated media. Optical breakdown, however, has to date been exclusively associated with compressive stress. The authors show that this is appropriate for water, but not for tissues for which the elastic-plastic material response needs to be considered. The acoustic transients following optical breakdown in water and cornea were measured with a fast hydrophone and the cavitation bubble dynamics, which is closely linked to the stress wave generation, was documented by flash photography. Breakdown in water produced amore » monopolar acoustic signal and a bubble oscillation in which the expansion and collapse phases were symmetric. Breakdown in cornea produced a bipolar acoustic signal coupled with a pronounced shortening of the bubble expansion phase and a considerable prolongation of its collapse phase. The tensile stress wave is related to the abrupt end of the bubble expansion. Numerical simulations using the MESA-2D code were performed assuming elastic-plastic material behavior in a wide range of values for the shear modulus and yield strength. The calculations revealed that consideration of the elastic-plastic material response is essential to reproduce the experimentally observed bipolar stress waves. The tensile stress evolves during the outward propagation of the acoustic transient and reaches an amplitude of 30--40% of the compressive pulse.« less

LRIG1 inhibits STAT3-dependent inflammation to maintain corneal homeostasis

PubMed Central

Nakamura, Takahiro; Hamuro, Junji; Takaishi, Mikiro; Simmons, Szandor; Maruyama, Kazuichi; Zaffalon, Andrea; Bentley, Adam J.; Kawasaki, Satoshi; Nagata-Takaoka, Maho; Fullwood, Nigel J.; Itami, Satoshi; Sano, Shigetoshi; Ishii, Masaru; Barrandon, Yann; Kinoshita, Shigeru

2013-01-01

Corneal integrity and transparency are indispensable for good vision. Cornea homeostasis is entirely dependent upon corneal stem cells, which are required for complex wound-healing processes that restore corneal integrity following epithelial damage. Here, we found that leucine-rich repeats and immunoglobulin-like domains 1 (LRIG1) is highly expressed in the human holoclone-type corneal epithelial stem cell population and sporadically expressed in the basal cells of ocular-surface epithelium. In murine models, LRIG1 regulated corneal epithelial cell fate during wound repair. Deletion of Lrig1 resulted in impaired stem cell recruitment following injury and promoted a cell-fate switch from transparent epithelium to keratinized skin-like epidermis, which led to corneal blindness. In addition, we determined that LRIG1 is a negative regulator of the STAT3-dependent inflammatory pathway. Inhibition of STAT3 in corneas of Lrig1–/– mice rescued pathological phenotypes and prevented corneal opacity. Additionally, transgenic mice that expressed a constitutively active form of STAT3 in the corneal epithelium had abnormal features, including corneal plaques and neovascularization similar to that found in Lrig1–/– mice. Bone marrow chimera experiments indicated that LRIG1 also coordinates the function of bone marrow–derived inflammatory cells. Together, our data indicate that LRIG1 orchestrates corneal-tissue transparency and cell fate during repair, and identify LRIG1 as a key regulator of tissue homeostasis. PMID:24316976

Frozen cultured sheets of epidermal keratinocytes in reepithelialization and repair of the cornea after photorefractive keratectomy.

PubMed

Castro-Muñozledo, Federico; Ozorno-Zarate, Jorge; Naranjo-Tackman, Ramon; Kuri-Harcuch, Walid

2002-09-01

To determine whether frozen cultured sheets of human allogeneic epidermal keratinocytes (CEAK) improved wound repair after experimental corneal ablation by photorefractive keratectomy (PRK). Hospital "Luis Sanchez Bulnes" de la Asociación para Evitar la Ceguera en Mexico, I.A.P, and Department of Cell Biology, CINVESTAV-IPN, Mexico City, Mexico. Transepithelial PRK was performed in the right eye of male albino rabbits to obtain a 112 microm deep and 6.0 mm diameter ablation zone. In 17 eyes, the ablations were covered with frozen CEAK; in 11 eyes, the ablations were covered with a disposable contact lens without the cultured sheets; and in the control group (13 eyes), the ablations were not covered. Subepithelial fibrosis and reepithelialization of the ablated zone were evaluated in serial paraffin-embedded tissue sections from all wounds. Treatment with CEAK reduced fibroblast proliferation and the inflammatory response beneath the ablated zone and produced better organization of the newly formed epithelium by eliminating significant hyperplasia or discontinuities in the periodic acid Shiff-stained basement membrane. It also led to accelerated reepithelialization. The use of frozen CEAK as a biologically active wound dressing improved tissue repair at 1 month in corneas ablated by transepithelial PRK in the male albino rabbit model. Treatment with CEAK could improve the outcome of PRK in humans.

Chronic Actinic Keratopathy—A Condition Associated with Conjunctival Elastosis (Pingueculae) and Typified by Characteristic Extracellular Concretions

PubMed Central

Klintworth, Gordon K.

1972-01-01

Morphologic observations on a peculiar type of corneal reaction with a predisposition for the superficial stroma of the interpalpebral portion of the cornea are reviewed. Histochemical evidence is provided which indicates that the corneal concretions, though not homogenous, are proteinaceous in nature and contain amino acids not normally detectable in the cornea. The corneal concretions were associated with conjunctival elastosis (pingueculae) in all 22 instances in which the eyes were sectioned in the horizontal plane. Identical concretions were identified within these associated pingueculae, as well as in a large percentage of other pingueculae and cutaneous lesions with actinic elastosis. The findings suggest that the abnormal material arises in the pericorneal conjunctival connective tissue from whence it diffuses into, and deposits in, the superficial corneal stroma. The data also raise the possibility that the concretions may be derived, at least in part, from altered elastic tissue. Morphologic and epidemiologic observations on the condition taken together strongly suggest that this unique reaction is a sequel to the cumulative effect of chronic actinic irradiation. Further observations on this keratopathy are needed to establish whether this unique response can be provoked by other noxious stimuli. ImagesFig 5Fig 6Fig 7Fig 8Fig 9Fig 10Fig 11Fig 12Fig 1Fig 2Fig 3Fig 4Fig 13Fig 14Fig 15 PMID:5021106

Topical administration of orbital fat-derived stem cells promotes corneal tissue regeneration.

PubMed

Lin, Ko-Jo; Loi, Mei-Xue; Lien, Gi-Shih; Cheng, Chieh-Feng; Pao, Hsiang-Yin; Chang, Yun-Chuang; Ji, Andrea Tung-Qian; Ho, Jennifer Hui-Chun

2013-06-14

Topical administration of eye drops is the major route for drug delivery to the cornea. Orbital fat-derived stem cells (OFSCs) possess an in vitro corneal epithelial differentiation capacity. Both the safety and immunomodulatory ability of systemic OFSC transplantation were demonstrated in our previous work. In this study, we investigated the safety, therapeutic effect, and mechanism(s) of topical OFSC administration in an extensive alkali-induced corneal wound. Corneal injury was created by contact of a piece of 0.5 N NaOH-containing filter paper on the corneal surface of a male Balb/c mouse for 30 s. The area of the filter paper covered the central 70% or 100% of the corneal surface. OFSCs (2 × 10(5)) in 5 μl phosphate-buffered saline (PBS) were given by topical administration (T) twice a day or by two intralimbal (IL) injections in the right cornea, while 5 μl of PBS in the left cornea served as the control. Topical OFSCs promoted corneal re-epithelialization of both the limbal-sparing and limbal-involved corneal wounds. In the first three days, topical OFSCs significantly reduced alkali-induced corneal edema and stromal infiltration according to a histopathological examination. Immunohistochemistry and immunofluorescence staining revealed that transplanted cells were easily detectable in the corneal epithelium, limbal epithelium and stroma, but only some of transplanted cells at the limbal epithelium had differentiated into cytokeratin 3-expressing cells. OFSCs did not alter neutrophil (Ly6G) levels in the cornea, but significantly reduced macrophage (CD68) infiltration and inducible nitrous oxide synthetase (iNOS) production during acute corneal injury as quantified by a Western blot analysis. Continuous topical administration of OFSCs for seven days improved corneal transparency, and this was accompanied by diffuse stromal engraftment of transplanted cells and differentiation into p63-expressing cells at the limbal area. The therapeutic effect of the topical administration of OFSCs was superior to that of the IL injection. OFSCs from the IL injection clustered in the limbal area and central corneal epithelium, which was associated with a persistent corneal haze. Topical OFSC administration is a simple, non-surgical route for stem cell delivery to promote corneal tissue regeneration through ameliorating acute inflammation and corneal epithelial differentiation. The limbal area serves as a niche for OFSCs differentiating into corneal epithelial cells in the first week, while the stroma is a potential site for anti-inflammation of OFSCs. Inhibition of corneal inflammation is related to corneal transparency.

Topical administration of orbital fat-derived stem cells promotes corneal tissue regeneration

PubMed Central

2013-01-01

Introduction Topical administration of eye drops is the major route for drug delivery to the cornea. Orbital fat-derived stem cells (OFSCs) possess an in vitro corneal epithelial differentiation capacity. Both the safety and immunomodulatory ability of systemic OFSC transplantation were demonstrated in our previous work. In this study, we investigated the safety, therapeutic effect, and mechanism(s) of topical OFSC administration in an extensive alkali-induced corneal wound. Methods Corneal injury was created by contact of a piece of 0.5 N NaOH-containing filter paper on the corneal surface of a male Balb/c mouse for 30 s. The area of the filter paper covered the central 70% or 100% of the corneal surface. OFSCs (2 × 105) in 5 μl phosphate-buffered saline (PBS) were given by topical administration (T) twice a day or by two intralimbal (IL) injections in the right cornea, while 5 μl of PBS in the left cornea served as the control. Results Topical OFSCs promoted corneal re-epithelialization of both the limbal-sparing and limbal-involved corneal wounds. In the first three days, topical OFSCs significantly reduced alkali-induced corneal edema and stromal infiltration according to a histopathological examination. Immunohistochemistry and immunofluorescence staining revealed that transplanted cells were easily detectable in the corneal epithelium, limbal epithelium and stroma, but only some of transplanted cells at the limbal epithelium had differentiated into cytokeratin 3-expressing cells. OFSCs did not alter neutrophil (Ly6G) levels in the cornea, but significantly reduced macrophage (CD68) infiltration and inducible nitrous oxide synthetase (iNOS) production during acute corneal injury as quantified by a Western blot analysis. Continuous topical administration of OFSCs for seven days improved corneal transparency, and this was accompanied by diffuse stromal engraftment of transplanted cells and differentiation into p63-expressing cells at the limbal area. The therapeutic effect of the topical administration of OFSCs was superior to that of the IL injection. OFSCs from the IL injection clustered in the limbal area and central corneal epithelium, which was associated with a persistent corneal haze. Conclusions Topical OFSC administration is a simple, non-surgical route for stem cell delivery to promote corneal tissue regeneration through ameliorating acute inflammation and corneal epithelial differentiation. The limbal area serves as a niche for OFSCs differentiating into corneal epithelial cells in the first week, while the stroma is a potential site for anti-inflammation of OFSCs. Inhibition of corneal inflammation is related to corneal transparency. PMID:23769140

Simulation model of an eyeball based on finite element analysis on a supercomputer

PubMed Central

Uchio, E.; Ohno, S.; Kudoh, J.; Aoki, K.; Kisielewicz, L. T.

1999-01-01

BACKGROUND/AIMS—A simulation model of the human eye was developed. It was applied to the determination of the physical and mechanical conditions of impacting foreign bodies causing intraocular foreign body (IOFB) injuries.
METHODS—Modules of the Hypermesh (Altair Engineering, Tokyo, Japan) were used for solid modelling, geometric construction, and finite element mesh creation based on information obtained from cadaver eyes. The simulations were solved by a supercomputer using the finite element analysis (FEA) program PAM-CRASH (Nihon ESI, Tokyo, Japan). It was assumed that rupture occurs at a strain of 18.0% in the cornea and 6.8% in the sclera and at a stress of 9.4 MPa for both cornea and sclera. Blunt-shaped missiles were shot and set to impact on the surface of the cornea or sclera at velocities of 30 and 60 m/s, respectively.
RESULTS—According to the simulation, the sizes of missile above which corneal rupture occurred at velocities of 30 and 60 m/s were 1.95 and 0.82 mm. The missile sizes causing scleral rupture were 0.95 and 0.75 mm at velocities of 30 and 60 m/s.
CONCLUSIONS—These results suggest that this FEA model has potential usefulness as a simulation tool for ocular injury and it may provide useful information for developing protective measures against industrial and traffic ocular injuries.

 PMID:10502567

Cost Minimization Analysis of Precut Cornea Grafts in Descemet Stripping Automated Endothelial Keratoplasty

PubMed Central

Yong, Kai-Ling; Nguyen, Hai V.; Cajucom-Uy, Howard Y.; Foo, Valencia; Tan, Donald; Finkelstein, Eric A.; Mehta, Jodhbir S.

2016-01-01

Abstract Descemet stripping automated endothelial keratoplasty (DSAEK) is the most common corneal transplant procedure. A key step in the procedure is preparing the donor cornea for transplantation. This can be accomplished via 1 of 3 alternatives: surgeon cuts the cornea on the day of surgery, the cornea is precut ahead of time in an offsite facility by a trained technician, or a precut cornea is purchased from an eye bank. Currently, there is little evidence on the costs and effectiveness of these 3 strategies to allow healthcare providers decide upon the preferred method to prepare grafts. The aim of this study was to compare the costs and relative effectiveness of each strategy. The Singapore National Eye Centre and Singapore Eye Bank performed both precut cornea and surgeon-cut cornea transplant services between 2009 and 2013. This study included 110 subjects who received precut cornea and 140 who received surgeon-cut cornea. Clinical outcomes and surgical duration were compared across the strategies using the propensity score matching. The cost of each strategy was estimated using the microcosting and consisted of facility costs and procedural costs including surgical duration. One-way sensitivity analysis and threshold analysis were performed. The cost for DSAEK was highest for the surgeon-cut approach ($13,965 per procedure), followed by purchasing precut corneas ($12,659) and then setting up precutting ($12,421). The higher procedural cost of the surgeon-cut approach was largely due to the longer duration of the procedure (surgeon-cut = 72.54 minutes, precut = 59.45 minutes, P < 0.001) and the higher surgeon fees. There was no evidence of differences in clinical outcomes between grafts that were precut or surgeon-cut. Threshold analysis demonstrated that if the number of cases was below 31 a year, the strategy that yielded the lowest cost was purchasing precut cornea from eye bank. If there were more than 290 cases annually, the cheapest option would be to setup precutting facility. Our findings suggest that it is more efficient for centers that are performing a large number of cornea transplants (more than 290 cases) to set up their own facility to conduct precutting. PMID:26937927

Comparison of the characteristics in hen and quail corneas as experimental models of refractive surgery.

PubMed

Gonçalves, G C; Pérez-Merino, P; Martínez-García, M C; Barcía, A; Merayo-Loves, J

2016-07-01

To compare the histological, morphological and the biophysical measurements between hen and quail corneas, in order to determine which of them were better suited for use as an animal model for research into corneal refractive surgery. A study was performed using the biophysical measurements of the cornea (curvature, thickness, refraction, and axial length) of 20 animals (10 hens and 10 quails). The corneas were then prepared for histological analysis under microscopy light. The analysis showed that both groups have the same number of corneal layers as the human cornea and with an evident Bowman's layer. The thickness of the hen cornea and axial length of the eye, 225.3±18.4μm and 12.8±0.25mm, respectively, were larger than that of the quail (P<.01 and P<.001, respectively). The radius of curvature for the hen central cornea, 3.65±0.08mm, was greater than that for the quail (P<.001), but the refractive power of each cornea was similar. The proportion of total corneal thickness of the hen stroma, 82.6%, was more similar to that of the human than was the quail stroma, 72.5%. Within the hen stroma, the density of keratocytes, 8.57±1.49 per 5,000μm(2), was about half that in the quail stroma (P<.005). Because of the large size of the hen cornea, the stromal thickness and proportional similarity of the corneal layers with human cornea, the hen maybe better than the quail as an alternative species suitable for use in studies of corneal refractive surgery. Copyright © 2016 Sociedad Española de Oftalmología. Published by Elsevier España, S.L.U. All rights reserved.

Regenerating Eye Tissues to Preserve and Restore Vision.

PubMed

Stern, Jeffrey H; Tian, Yangzi; Funderburgh, James; Pellegrini, Graziella; Zhang, Kang; Goldberg, Jeffrey L; Ali, Robin R; Young, Michael; Xie, Yubing; Temple, Sally

2018-06-01

Ocular regenerative therapies are on track to revolutionize treatment of numerous blinding disorders, including corneal disease, cataract, glaucoma, retinitis pigmentosa, and age-related macular degeneration. A variety of transplantable products, delivered as cell suspensions or as preformed 3D structures combining cells and natural or artificial substrates, are in the pipeline. Here we review the status of clinical and preclinical studies for stem cell-based repair, covering key eye tissues from front to back, from cornea to retina, and including bioengineering approaches that advance cell product manufacturing. While recognizing the challenges, we look forward to a deep portfolio of sight-restoring, stem cell-based medicine. VIDEO ABSTRACT. Copyright © 2018 Elsevier Inc. All rights reserved.

Glycerol dehydration of native and diabetic animal tissues studied by THz-TDS and NMR methods

PubMed Central

Smolyanskaya, O. A.; Schelkanova, I. J.; Kulya, M. S.; Odlyanitskiy, E. L.; Goryachev, I. S.; Tcypkin, A. N.; Grachev, Ya. V.; Toropova, Ya. G.; Tuchin, V. V.

2018-01-01

The optical clearing method has been widely used for different spectral ranges where it provides tissue transparency. In this work, we observed the enhanced penetration of the terahertz waves inside biological samples (skin, kidney, and cornea) treated with glycerol solutions inducing changes of optical and dielectric properties. It was supported by the observed trend of free-to-bound water ratio measured by the nuclear magnetic resonance (NMR) method. The terahertz clearing efficiency was found to be less for diabetic samples than for normal ones. Results of the numerical simulation proved that pulse deformation is due to bigger penetration depth caused by the reduction of absorption and refraction at optical clearing. PMID:29541513

Quantification of the ultraviolet radiation (UVR) field in the human eye in vivo using novel instrumentation and the potential benefits of UVR blocking hydrogel contact lens.

PubMed

Walsh, J E; Bergmanson, J P; Wallace, D; Saldana, G; Dempsey, H; McEvoy, H; Collum, L M

2001-09-01

Certain degenerative eye conditions occur predominantly nasally, at the limbal region, and are associated with solar ultraviolet radiation (UVR) induced damage. The relative contribution to the in vivo ocular flux of (a) the reflection of UVR incident on the skin of the nose onto the nasal limbus, and (b) the focusing of UVR incident on the temporal side of the cornea onto the nasal limbus were examined. A novel photodiode sensor array was used to measure the UVR field across the eye. In addition, a novel spectrometer set-up was used to measure the spectrum of radiation refracted across the cornea. The efficacy of UVR blocking hydrogel contact lenses in filtering incident UVR was assessed in vivo. Qualitative and quantitative data indicated an increase nasally of UVR. Photodiode readings showed a net UVR increase from the temporal to the nasal side. Transmission curves showed that most UVR incident on the limbal region is either absorbed by, or transmitted through, the ocular tissues. This radiation is filtered by UVR blocking soft contact lens. An increased UVR flux on the nasal side of the eye, due to reflection off the nasal skin, was identified in vivo. Any UVR passing through the cornea is either absorbed by the conjunctiva and/or transmitted through it onto the sclera where it is absorbed. UVR blocking hydrogel contact lenses can eliminate these sources of UVR.

Influence of drug loading and type of ointment base on the in vitro performance of acyclovir ophthalmic ointment.

PubMed

Al-Ghabeish, Manar; Xu, Xiaoming; Krishnaiah, Yellela S R; Rahman, Ziyaur; Yang, Yang; Khan, Mansoor A

2015-11-30

The availability of in vitro performance tests such as in vitro drug release testing (IVRT) and in vitro permeation testing (IVPT) are critical to comprehensively assure consistent delivery of the active component(s) from semisolid ophthalmic drug products. The objective was to study the impact of drug loading and type of ointment base on the in vitro performance (IVRT and IVPT) of ophthalmic ointments using acyclovir as a model drug candidate. The in vitro drug release for the ointments was evaluated using a modified USP apparatus 2 with Enhancer cells. The transcorneal permeation was carried out using rabbit cornea on modified vertical Franz cells. The drug retention in cornea (DRC) was also determined at the end of transcorneal drug permeation study. The in vitro drug release, transcorneal drug permeation as well as DRC exhibited a proportional increase with increasing drug loading in the ointment. On comparing the in vitro drug release profile with transcorneal permeation profile, it appears that drug release from the ointment is controlling acyclovir transport through the cornea. Furthermore, enhanced in vitro transcorneal permeation relative to the in vitro drug release underscores the importance of the interplay between the physiology of the ocular tissue and ointment formulation. The results indicated that IVRT and IVPT could be used to discriminate the impact of changes in drug load and formulation composition of ophthalmic ointments. Copyright © 2015. Published by Elsevier B.V.

Measuring ocular characteristics after gel injection adjustable keratoplasty (GIAK) in the rabbit

NASA Astrophysics Data System (ADS)

Comander, Jason I.; Parel, Jean-Marie A.; Simon, Gabriel; Takesue, Yoshiko; Villain, Franck L.

1995-05-01

Gel Injection Adjustable Keratoplasty (GIAK) is a refractive surgery procedure which uses an ocular ring implant made of a polyethylene oxide hydrogel to cause a refractive change in the cornea. Unlike laser photo refractive keratectomy, GIAK does not interfere with the central cornea because the ring lies around the optical axis. Thus, vision can be assessed immediately after surgery. Our in vivo study was designed to quantify GIAK's effects on tissues, the biocompatibility of the polymer and in the process investigate which ocular changes in the rabbit model can be monitored with precision using current technology. Thirty-two young rabbits underwent a delamination in one eye, 22 of which were injected with a new polymeric gel. Corneal topography, keratometry, pachymetry, and tonometry were performed on both eyes for up to 105 days. All corneas flattened with growth. In GIAK animals, we found an average flattening of 6.51 +/- 1.23 diopters (p < 0.0001) relative to the fellow eye. No statistically significant regression over the 102 days was observed. Intraocular pressure dropped slightly by 0.69 +/- 1.21 mmHg (p equals 0.025), a clinically insignificant value, while no significant change was detected in corneal thickness. Keratometry can be tracked in rabbits after GIAK surgery from POD 1. Measuring unoperated fellow eyes allows for the effects of surgery to be assessed without bias from growth. Using this protocol, GIAK was shown to be stable. It was more difficult to draw conclusions from pachymetry, tonometry, and topography data.

Protocol for vital dye staining of corneal endothelial cells.

PubMed

Park, Sunju; Fong, Alan G; Cho, Hyung; Zhang, Cheng; Gritz, David C; Mian, Gibran; Herzlich, Alexandra A; Gore, Patrick; Morganti, Ashley; Chuck, Roy S

2012-12-01

To describe a step-by-step methodology to establish a reproducible staining protocol for the evaluation of human corneal endothelial cells. Four procedures were performed to determine the best protocol. (1) To determine the optimal trypan blue staining method, goat corneas were stained with 4 dilutions of trypan blue (0.4%, 0.2%, 0.1%, and 0.05%) and 1% alizarin red. (2) To determine the optimal alizarin red staining method, goat corneas were stained with 2 dilutions of alizarin red (1% and 0.5%) and 0.2% trypan blue. (3) To ensure that trypan blue truly stains damaged cells, goat corneas were exposed to either 3% hydrogen peroxide or to balanced salt solution, and then stained with 0.2% trypan blue and 0.5% alizarin red. (4) Finally, fresh human corneal buttons were examined; 1 group was stained with 0.2% trypan blue and another group with 0.4% trypan blue. For the 4 procedures performed, the results are as follows: (1) trypan blue staining was not observed in any of the normal corneal samples; (2) 0.5% alizarin red demonstrated sharper cell borders than 1% alizarin red; (3) positive trypan blue staining was observed in the hydrogen peroxide exposed tissue in damaged areas; (4) 0.4% trypan blue showed more distinct positive staining than 0.2% trypan blue. We were able to determine the optimal vital dye staining conditions for human corneal endothelial cells using 0.4% trypan blue and 0.5% alizarin red.

Identification of biomechanical properties of the cornea: the ocular response analyzer.

PubMed

Terai, Naim; Raiskup, Frederik; Haustein, Michael; Pillunat, Lutz E; Spoerl, Eberhard

2012-07-01

Several methods have been devised for measuring geometric parameters of the cornea but, until now, the biomechanics of the cornea have been largely ignored. The relatively new Ocular Response Analyzer (ORA) provides such biomechanical information. In order to correctly interpret the underlying biomechanics of ORA data, we review reported ORA measurements and provide a compendium of factors influencing these measurements, with discussion of possible explanations for ORA measurement results. This review comprised a literature search using "ocular response analyzer" and "ocular response analyser" as keywords. We reviewed and compared reported results from recent ORA studies so obtained, with an eye to understanding corneal biomechanics. Several ORA biomechanical parameters of the cornea - corneal hysteresis (CH) and corneal resistant factor (CRF) - characterize the viscoelastic properties of the cornea, especially those of the ground substance. The impact on CH and CRF values of various independent factors, e.g. intraocular pressure (IOP), age, central corneal thickness (CCT), and corneal swelling, are discussed. The impact on CH and CRF of treatment-related structural changes of the cornea, i.e. those occurring after refractive surgical procedures, placement of intracorneal rings, and collagen crosslinking (CXL), as well as pathological changes of the cornea, e.g. those resulting from keratoconus, edema, and glaucoma, are discussed. Changes in CRF and CH may be reflective of structural changes in the ground substance of the cornea. Thus, ORA provides invaluable information for delineating biomechanical conditions pertaining to the cornea, with special regard to ocular diseases, e.g. keratoconus and glaucoma.

Comparison of confocal microscopy and two-photon microscopy in mouse cornea in vivo.

PubMed

Lee, Jun Ho; Lee, Seunghun; Gho, Yong Song; Song, In Seok; Tchah, Hungwon; Kim, Myoung Joon; Kim, Ki Hean

2015-03-01

High-resolution imaging of the cornea is important for studying corneal diseases at cellular levels. Confocal microscopy (CM) has been widely used in the clinic, and two-photon microscopy (TPM) has recently been introduced in various pre-clinical studies. We compared the performance of CM and TPM in normal mouse corneas and neovascularized mouse corneas induced by suturing. Balb/C mice and C57BL/6 mice expressing green fluorescent protein (GFP) were used to compare modalities based on intrinsic contrast and extrinsic fluorescence contrast. CM based on reflection (CMR), CM based on fluorescence (CMF), and TPM based on intrinsic/extrinsic fluorescence and second harmonic generation (SHG) were compared by imaging the same sections of mouse corneas sequentially in vivo. In normal mouse corneas, CMR visualized corneal cell morphologies with some background noise, and CMF visualized GFP expressing corneal cells clearly. TPM visualized corneal cells and collagen in the stroma based on fluorescence and SHG, respectively. However, in neovascularized mouse corneas, CMR could not resolve cells deep inside the cornea due to high background noise from the effects of increased structural irregularity induced by suturing. CMF and TPM visualized cells and induced vasculature better than CMR because both collect signals from fluorescent cells only. Both CMF and TPM had signal decays with depth due to the structural irregularity, with CMF having faster signal decay than TPM. CMR, CMF, and TPM showed different degrees of image degradation in neovascularized mouse corneas. Copyright © 2015 Elsevier Ltd. All rights reserved.

Femtosecond laser cutting of multiple thin corneal stromal lamellae for endothelial bioengineering.

PubMed

Bernard, Aurélien; He, Zhiguo; Forest, Fabien; Gauthier, Anne-Sophie; Peocʼh, Michel; Dumollard, Jean-Marc; Acquart, Sophie; Montard, Romain; Delbosc, Bernard; Gain, Philippe; Thuret, Gilles

2015-02-01

To assess the feasibility of cutting multiple thin stromal lamellae in human donor corneas using a commercial femtosecond laser (FSL) to provide cell carriers for future endothelial graft bioengineering. Eight edematous organ-cultured corneas not suitable for grafting for endothelial reasons were mounted on a Ziemer anterior chamber and cut with a Z6 FSL with 6 successive parallel cuts, from depth to surface. Target thickness of each lamella ranged from 100 to 150 μm depending on initial corneal thickness. Thickness was measured using anterior segment optical coherence tomography before and after cutting on mounted corneas, and on each stromal lamella after detachment. Scanning electron microscopy observation was performed on 4 lamellae and histological cross sections on 1 cornea before detachment. A median of 5 (minimum 3, maximum 7) lamellae was obtained per cornea. All lamellae still attached were the most posterior ones, suggesting that FSL was less efficient because of light scattering by edematous stroma. Cut precision and postdetachment swelling were correlated with anterior-posterior position within the cornea. Median lamella thickness was 127 μm (56-222 μm) before detachment and 196 μm (80-304 μm) after detachment. Surface state was consistent with previously reported FSL lamellar cuts during Descemet stripping automated endothelial keratoplasty. Up to 7 thin lamellae can be cut in stored corneas with an FSL. This method, once optimized primarily by using deswelled, more transparent corneas, could prove effective for recycling unsuitable donor corneas in corneal bioengineering processes.

Analysis of the scattering performance of human retinal tissue layers

NASA Astrophysics Data System (ADS)

Zhu, Dan; Gao, Zhisan; Ye, Haishui; Yuan, Qun

2017-02-01

Human retina is different from other ocular tissues, such as cornea, crystalline lens and vitreous because of high scattering performance. As an anisotropic tissue, we cannot neglect its impact on the polarization state of the scattered light. In this paper, Mie scattering and radiative transfer theory are applied to analyze the polarization state of backscattered light from four types of retinal tissues, including neural retina, retinal pigment epithelial (RPE), choroid and sclera. The results show that the most backscattered zones in different depths have almost the same electrical fields of Jones vector, which represents the polarization state of light, whether neural retina layer is under normal incidence or oblique incidence. Very little change occurs in the polarization of backscattered light compared to that of the incident light. Polarization distribution of backward scattered light from neural retina layer doesn't make apparent effects on polarization phase shifting in spectral domain OCT because its thickness is far less than photon mean free path, while other retinal tissues do not meet this rule.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ediger, M.N.

The laser-induced fluorescence spectrum of rabbit cornea irradiated at ablative intensities was measured. This system directly measured the radiant exposure of fluorescence transmitted through the cornea when the anterior surface of the cornea was irradiated by an ArF excimer laser. Evidence of changing spectral characteristics as a function of total laser dose suggests photochemical changes in the cornea may be occurring. Results are compared with previous data of laser-induced fluorescence in other models and detection schemes.

Collagen organization in the chicken cornea and structural alterations in the retinopathy, globe enlarged (rge) phenotype--an X-ray diffraction study.

PubMed

Boote, Craig; Hayes, Sally; Jones, Simon; Quantock, Andrew J; Hocking, Paul M; Inglehearn, Chris F; Ali, Manir; Meek, Keith M

2008-01-01

An investigation into the collagenous structure of the mature avian cornea is presented. Wide-angle X-ray diffraction is employed to assess collagen organization in 9-month-old chicken corneas. The central 2-4mm corneal region features a preponderance of fibrils directed along the superior-inferior and nasal-temporal orthogonal meridians. More peripherally the orientation of fibrils alters in favor of a predominantly tangential arrangement. The chicken cornea appears to be circumscribed by an annulus of fibrils that extends into the limbus. The natural arrangement of collagen in the chicken cornea is discussed in relation to corneal shape and the mechanical requirements of avian corneal accommodation. Equivalent data are also presented from age-matched blind chickens affected with the retinopathy, globe enlarged (rge) mutation, characterized by an abnormally thick and flat cornea. The data indicate considerable realignment and redistribution of collagen lamellae in the peripheral rge cornea. In contrast to normal chickens, no obvious tangential collagen alignment was evident in the periphery of rge corneas. In mammals, the presence of a limbal fibril annulus is believed to be important in corneal shape preservation. We postulate that corneal flattening in rge chickens may be related to biomechanical changes brought about by an alteration in collagen arrangement at the corneal periphery.

Role of solvent-mediated carbodiimide cross-linking in fabrication of electrospun gelatin nanofibrous membranes as ophthalmic biomaterials.

PubMed

Chou, Shih-Feng; Luo, Li-Jyuan; Lai, Jui-Yang; Ma, David Hui-Kang

2017-02-01

Due to their ability to mimic the structure of extracellular matrix, electrospun gelatin nanofibers are promising cell scaffolding materials for tissue engineering applications. However, the hydrophilic gelatin molecules usually need stabilization before use in aqueous physiological environment. Considering that biomaterials cross-linked via film immersion technique may have a more homogeneous cross-linked structure than vapor phase cross-linking, this work aims to investigate the chemical modification of electrospun gelatin nanofibrous membranes by liquid phase carbodiimide in the presence of ethanol/water co-solvents with varying ethanol concentrations ranging from 80 to 99.5vol%. The results of characterization showed that increasing water content in the binary reaction solvent system increases the extent of cross-linking of gelatin nanofibers, but simultaneously promotes the effect of biopolymer swelling and distortion in fiber mat structure. As compared to non-cross-linked counterparts, carbodiimide treated gelatin nanofibrous mats exhibited better thermal and biological stability where the shrinkage temperature and resistance to enzymatic degradation varied in response to ethanol/water solvent composition-mediated generation of cross-links. Irrespective of their cross-linking density, all studied membrane samples did not induce any responses in ocular epithelial cell cultures derived from cornea, lens, and retina. Unlike many other cross-linking agents and/or methods (e.g., excessive vapor phase cross-linking) that may pose a risk of toxicity, our study demonstrated that these nanofibrous materials are well tolerated by anterior segment tissues. These findings also indicate the safety of using ethanol/water co-solvents for chemical cross-linking of gelatin to engineer nanofibrous materials with negligible biological effects. In summary, the present results suggest the importance of solvent-mediated carbodiimide cross-linking in modulating structure-property relationship without compromising in vitro and in vivo biocompatibility of electrospun gelatin nanofibers for future ophthalmic applications. Copyright © 2016 Elsevier B.V. All rights reserved.

Intrastromal Injection of China Painting Ink in Corneas of Male Rabbits: Clinical and Histological Study.

PubMed

Alsmman Hassan, Alahmady Hamad; Abd Elhaliem Soliman, Nesreen Gamal-Eldeen

2016-01-01

Background. Many patients with corneal opacity or complicated cataract in blind eye ask for cosmoses. In this study we tried to investigate the staining of corneas of male rabbits by Rotring China painting ink and to study the histological changes. Method. 10 eyes of 10 male Baladi Egyptian rabbits were injected (0.1 mL) intrastromally in the cornea by the use of China painting ink (Rotring Tinta China) through insulin syringe (27-gauge needle) by single injection; clinical follow-up is for 6 months and lastly the rabbits were scarified and the stained eyes were enucleated for histological analysis. Results. Clinically the stain was stable in color and distribution in corneas with no major complications. Histological results of the stained rabbit corneas showed blackish pigmentation in the corneal stroma without any inflammatory cellular infiltration. Some fibroblast cells had pigment granules in their cytoplasm in the adjacent layers. Conclusion. Corneal staining by China painting ink is effective and safe in staining of male rabbits cornea; however further study in human corneas with longer follow-up period is advisable.

Dual interferometer for dynamic measurement of corneal topography

NASA Astrophysics Data System (ADS)

Micali, Jason D.; Greivenkamp, John E.

2016-08-01

The cornea is the anterior most surface of the eye and plays a critical role in vision. A thin fluid layer, the tear film, coats the outer surface of the cornea and serves to protect, nourish, and lubricate the cornea. At the same time, the tear film is responsible for creating a smooth continuous surface, where the majority of refraction takes place in the eye. A significant component of vision quality is determined by the shape of the cornea and stability of the tear film. A dual interferometer system for measuring the dynamic corneal topography is designed, built, verified, and qualified by testing on human subjects. The system consists of two coaligned simultaneous phase-shifting polarization-splitting Twyman-Green interferometers. The primary interferometer measures the surface of the tear film while the secondary interferometer tracks the absolute position of the cornea, which provides enough information to reconstruct the absolute shape of the cornea. The results are high-resolution and high-accuracy surface topography measurements of the in vivo tear film and cornea that are captured at standard camera frame rates.

Residual corneal stroma in big-bubble deep anterior lamellar keratoplasty: a histological study in eye-bank corneas.

PubMed

McKee, Hamish D; Irion, Luciane C D; Carley, Fiona M; Jhanji, Vishal; Brahma, Arun K

2011-10-01

To determine if residual corneal stroma remains on the recipient posterior lamella in big-bubble deep anterior lamellar keratoplasty (DALK). Pneumodissection using the big-bubble technique was carried out on eye-bank corneas mounted on an artificial anterior chamber. Samples that had a successful big-bubble formation were sent for histological evaluation to determine if any residual stroma remained on the Descemet membrane (DM). Big-bubble formation was achieved in 32 donor corneas. Two distinct types of big-bubble were seen: the bubble had either a white margin (30 corneas) or a clear margin (two corneas). The posterior lamellae of all the white margin corneas showed residual stroma on DM with a mean central thickness of 7.0 μm (range 2.6-17.4 μm). The clear margin corneas showed no residual stroma on DM. It should no longer be assumed that big-bubble DALK, where the bubble has a white margin, routinely bares DM. True baring of DM may only occur with the less commonly seen clear margin bubble.

Influence of the ablation plume on the removal process during ArF-excimer laser photoablation

NASA Astrophysics Data System (ADS)

Doerbecker, Christina; Lubatschowski, Holger; Lohmann, Stefan; Ruff, Christine; Kermani, Omid; Ertmer, Wolfgang

1996-01-01

Correction of myopia with the ArF-excimer laser (PRK) sometimes leads to a so called 'central island' formation on the anterior corneal surface. The attenuation of the laser beam by the ablation plume might be one reason for this phenomenon. The attenuation properties of the ablation plume were investigated by a probe beam parallel to the surface of the tissue probe. By varying the laser parameters (fluence, repetition rate, spot size) and the target tissue (cornea, PMMA) the attenuation of the probe beam was measured time and spatial resolved. As a result of this study, a significant influence of the removal process due to scattering and absorption within the ablation plume can be assumed as a function of repetition rate, spot size and air flow on the tissue surface.

Mid-IR laser system for advanced neurosurgery

NASA Astrophysics Data System (ADS)

Klosner, M.; Wu, C.; Heller, D. F.

2014-03-01

We present work on a laser system operating in the near- and mid-IR spectral regions, having output characteristics designed to be optimal for cutting various tissue types. We provide a brief overview of laser-tissue interactions and the importance of controlling certain properties of the light beam. We describe the principle of operation of the laser system, which is generally based on a wavelength-tunable alexandrite laser oscillator/amplifier, and multiple Raman conversion stages. This configuration provides robust access to the mid-IR spectral region at wavelengths, pulse energies, pulse durations, and repetition rates that are attractive for neurosurgical applications. We summarize results for ultra-precise selective cutting of nerve sheaths and retinas with little collateral damage; this has applications in procedures such as optic-nerve-sheath fenestration and possible spinal repair. We also report results for cutting cornea, and dermal tissues.

A minimal model of epithelial tissue dynamics and its application to the corneal epithelium

NASA Astrophysics Data System (ADS)

Henkes, Silke; Matoz-Fernandez, Daniel; Kostanjevec, Kaja; Coburn, Luke; Sknepnek, Rastko; Collinson, J. Martin; Martens, Kirsten

Epithelial cell sheets are characterized by a complex interplay of active drivers, including cell motility, cell division and extrusion. Here we construct a particle-based minimal model tissue with only division/death dynamics and show that it always corresponds to a liquid state with a single dynamic time scale set by the division rate, and that no glassy phase is possible. Building on this, we construct an in-silico model of the mammalian corneal epithelium as such a tissue confined to a hemisphere bordered by the limbal stem cell zone. With added cell motility dynamics we are able to explain the steady-state spiral migration on the cornea, including the central vortex defect, and quantitatively compare it to eyes obtained from mice that are X-inactivation mosaic for LacZ.

Angiogenesis and Therapeutic Approaches to NF1 Tumors

DTIC Science & Technology

2007-04-01

corneal neovascularization model was developed. In this model, the avascularity of the cornea highly facilitates the quantification of neovascularture...wild-type corneas in avascular area. However, in the NV zone, the number of macrophage was 4.6-fold greater in Nf1þ /– corneas than wild-type corneas...GEM tumor classification because of low cellularity and no necrosis . They exceed that clas- sification, however, due to their low to moderate prolif

DOE Office of Scientific and Technical Information (OSTI.GOV)

Larin, Kirill V; Ghosn, M G

The passive diffusion of drugs through the epithelial surfaces of an eye (the most widespread method for medical treatment of various diseases) is considered. The permeability of water and drugs through rabbit cornea was measured in the isolated cornea (separate from an eye) and in the whole cornea. The permeability coefficients of water and dexamethasone were estimated by the method of optical coherence tomography (OCT). Because multiple photon scattering introduces noise and distortions to the OCT signal, measurements were performed at depths up to 500 {mu}m where most likely single scattering of light occurs in cornea. It is shown thatmore » the permeability coefficients in the isolated and whole cornea strongly differ from each other. For example, the water permeability in the isolated and whole cornea is (7.09{+-}0.12)x10{sup -5} and (1.71{+-}0.51)x10{sup -5} cm s{sup -1}, respectively. (special issue devoted to multiple radiation scattering in random media)« less

Cosmetic Preservatives as Therapeutic Corneal and Scleral Tissue Cross-Linking Agents

PubMed Central

Babar, Natasha; Kim, MiJung; Cao, Kerry; Shimizu, Yukari; Kim, Su-Young; Takaoka, Anna; Trokel, Stephen L.; Paik, David C.

2015-01-01

Purpose. Previously, aliphatic β-nitroalcohols (BNAs) have been studied as a means to chemically induce tissue cross-linking (TXL) of cornea and sclera. There are a number of related and possibly more potent agents, known as formaldehyde releasers (FARs), that are in commercial use as preservatives in cosmetics and other personal care products. The present study was undertaken in order to screen such compounds for potential clinical utility as therapeutic TXL agents. Methods. A chemical registry of 62 FARs was created from a literature review and included characteristics relevant to TXL such as molecular weight, carcinogenicity/mutagenicity, toxicity, hydrophobicity, and commercial availability. From this registry, five compounds [diazolidinyl urea (DAU), imidazolidinyl urea (IMU), sodium hydroxymethylglycinate (SMG), DMDM hydantoin (DMDM), 5-Ethyl-3,7-dioxa-1-azabicyclo [3.3.0] octane (OCT)] were selected for efficacy screening using two independent systems, an ex vivo rabbit corneal cross-linking simulation setup and incubation of cut scleral tissue pieces. Treatments were conducted at pH 7.4 or 8.5 for 30 minutes. Efficacy was evaluated using thermal denaturation temperature (Tm), and cell toxicity was studied using the trypan blue exclusion method. Results. Cross-linking effects in the five selected FARs were pH and concentration dependent. Overall, the Tm shifts were in agreement with both cornea and sclera. By comparison with BNAs previously reported upon, the FARs identified in this study were significantly more potent but with similar or better cytotoxicity. Conclusions. The FARs, a class of compounds well known to the cosmetic industry, may have utility as therapeutic TXL agents. The compounds studied thus far show promise and will be further tested. PMID:25634979

Development of Methods for the Real-Time and Rapid Identification and Detection of TSE in Living Animals Using Fluorescence Spectroscopy of the Eye

DTIC Science & Technology

2006-07-01

retina . Our experiments have so far been limited to sheep. Our experiments have been designed to address the following questions: 1. Can ocular...We dissected sheep and cow eyes and performed fluorescence spectroscopy on all the major eye components and reports that the cornea, lens, retina ...excitation wavelengths λex = 410, 470, and 520 nm: retina ; optic nerve; outer tissue (sclera); and lens. • Contrary to the conclusions we

Early Intervention Stem Cell-Based Therapy (EISCBT) for Corneal Burns and Trauma

DTIC Science & Technology

2015-10-01

be held in place on the cornea by a soft contact lens. We will optimize means of storing the ReCoBand frozen so they will be available to doctors... regeneration in vivo Task 3. Assess plastic compressed collagen as a stem cell delivery vehicle Task 4. Assess cell sheets as delivery vehicle for stem...Stromal Stem Cells (CSSC) are obtained from biopsies of corneal tissue . The quality and potency of individual stem cell lines varies greatly from one

Micro-mechanical model for the tension-stabilized enzymatic degradation of collagen tissues

NASA Astrophysics Data System (ADS)

Nguyen, Thao; Ruberti, Jeffery

We present a study of how the collagen fiber structure influences the enzymatic degradation of collagen tissues. Experiments of collagen fibrils and tissues show that mechanical tension can slow and halt enzymatic degradation. Tissue-level experiments also show that degradation rate is minimum at a stretch level coincident with the onset of strain-stiffening in the stress response. To understand these phenomena, we developed a micro-mechanical model of a fibrous collagen tissue undergoing enzymatic degradation. Collagen fibers are described as sinusoidal elastica beams, and the tissue is described as a distribution of fibers. We assumed that the degradation reaction is inhibited by the axial strain energy of the crimped collagen fibers. The degradation rate law was calibrated to experiments on isolated single fibrils from bovine sclera. The fiber crimp and properties were fit to uniaxial tension tests of tissue strips. The fibril-level kinetic and tissue-level structural parameters were used to predict tissue-level degradation-induced creep rate under a constant applied force. We showed that we could accurately predict the degradation-induce creep rate of the pericardium and cornea once we accounted for differences in the fiber crimp structure and properties.

The role of crumbs genes in the vertebrate cornea.

PubMed

Beyer, Jill; Zhao, Xinping C; Yee, Richard; Khaliq, Shagufta; McMahon, Timothy T; Ying, Hongyu; Yue, Beatrice Y J T; Malicki, Jarema J

2010-09-01

To evaluate the role of crumbs genes and related epithelial polarity loci in the vertebrate cornea. The authors used histologic analysis and electron microscopy to evaluate the corneas of zebrafish mutant for a crumbs locus oko meduzy (ome) and in mutants of four other loci, nagie oko (nok), heart and soul (has), mosaic eyes (moe), and ncad (formerly glass onion), that function in the same or related genetic pathways. In parallel, they performed an evaluation of corneas in human carriers of a crumbs gene, CRB1, and mutations using topography and biomicroscopy. The expression of the CRB1 gene in the normal human cornea was examined by polymerase chain reaction (PCR) and immunohistochemical staining. The corneas of zebrafish mutants display severe abnormalities of the epithelial and stromal layers. The epithelial cells do not properly adhere to each other, and fluid-filled spaces form between them. In addition, the layering of the corneal stroma is poorly formed or absent. The corneas of human carriers of CRB1 mutations display shape deviations compared with what has been observed in normal individuals. A PCR product of the correct size was obtained from normal human corneal samples. Sequence analyses confirmed its identity to be the human CRB1 gene. Immunohistochemical staining using anti-CRB1 yielded positive brown deposits in the human cornea. crumbs genes play a role in the differentiation of the vertebrate cornea. Corneal defects associated with crumbs gene mutations are very severe in the zebrafish model and, in comparison, appear clinically less pronounced in the human eye.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cenedella, R.J.; Fleschner, C.R.

The authors developed a direct chemical approach for estimating the rate of turnover of the corneal epithelium in vivo. The method was used to examine the effects of lovastatin, a potent inhibitor of cholesterol biosynthesis, on proliferation and turnover of the epithelium. Corneal DNA was labeled by pulse injection (IP) of the rat with 3H-thymidine, and 3H-labeled DNA was recovered from peripheral and central corneas over the next 15 days. Only the epithelium became labeled, and the loss of label by cell desquamation began 3 days after injection. The loss of 3H-DNA from the cornea (peripheral plus central region) followedmore » first-order kinetics. The half-life of the disappearance was about 3 days. The peripheral cornea became more highly labeled than the central cornea and began to lose 3H-DNA before the central cornea. These observations support the possibility of a higher mitotic rate in the peripheral region and the centripetal movement of a population of peripheral epithelial cells in the normal cornea. The half-lives of the disappearance of 3H-DNA from peripheral and central corneas measured between days 5 and 15 postinjection were identical, both at 3 days. Complete turnover of the corneal epithelium would, therefore, require about 2 weeks (4-5 half-lives). Treatment of the rat with lovastatin had no obvious effects upon the proliferation or turnover of the corneal epithelium. Although lovastatin inhibited corneal 3-hydroxy-3-methylglutaryl coenzyme A reductase, the key regulatory enzyme of cholesterol synthesis, the cornea compensated by induction of this enzyme so that there was no net inhibition of cholesterol synthesis in the cornea.« less

Treatment of alkali-injured cornea by cyclosporine A-loaded electrospun nanofibers - An alternative mode of therapy.

PubMed

Cejkova, Jitka; Cejka, Cestmir; Trosan, Peter; Zajicova, Alena; Sykova, Eva; Holan, Vladimir

2016-06-01

In this study we tried to develop a new approach to suppress inflammation and neovascularization in the alkali-injured rabbit cornea. For this reason Cyclosporine A (CsA)-loaded electrospun nanofibers were transferred onto the ocular surface injured with alkali (0.25 N NaOH). Damaged corneas were divided into the following groups: untreated, treated with CsA eye drops, treated with nanofibers drug-free and treated with CsA-loaded nanofibers. Healthy rabbit corneas served as controls. Drug-free nanofibers and CsA-loaded nanofibers were transferred onto the damaged corneal surface immediately after the injury and sutured to conjunctiva. On day five after the injury the nanofibers were removed. The animals from all groups were sacrificed on day twelve after the injury. The extent of the inflammatory reaction and corneal healing were examined macroscopically, immunohistochemically and biochemically. The central corneal thickness was measured using an ultrasonic pachymeter. When compared with untreated injured corneas, injured corneas treated with drug-free nanofibers or injured corneas treated with CsA eye drops, the number of CD3-positive cells (T lymphocytes) and the production of pro-inflammatory cytokines were strongly reduced in corneas treated with CsA-loaded nanofibers, which was associated with the significantly decreased expression of matrix metalloproteinase 9, inducible nitric oxide synthase, vascular endothelial growth factor and active caspase-3. CsA-loaded nanofibers effectively suppressed corneal inflammation and corneal neovascularization. Central corneal thickness restored to levels before injury only in corneas treated with CsA-loaded nanofibers. Corneal transparency was highly restored in these corneas. It is suggested that the beneficial effect of CsA-loaded nanofibers was associated with the continuous release of CsA from nanofibers and continuous affection of damaged cornea by CsA. The suture of nanofibers to conjunctiva and the closed eyes contributed to beneficial corneal healing. This is in contrast to CsA eye drops, which are quickly washed from the ocular surface and the contact of CsA with the damaged cornea was limited. In conclusion, the approach with CsA-loaded nanofibers could represent an effective alternative mode of therapy for corneal chemical burns. Copyright © 2016 Elsevier Ltd. All rights reserved.

Transient expression of collagen type II at epitheliomesenchymal interfaces during morphogenesis of the cartilaginous neurocranium.

PubMed

Thorogood, P; Bee, J; von der Mark, K

1986-08-01

In the avian embryo a matrix-mediated tissue interaction between retinal pigmented epithelium and neural crest-derived periocular mesenchyme leads to the differentiation of (scleral) cartilage. The composition of the extracellular matrix at the interface between these two tissues has been examined immunohistochemically, both during and after the interaction has taken place. Of the matrix components studied (fibronectin, laminin, and collagen types I, II, IV, and V) only collagen type II displayed a dramatic change in distribution between the two stages. During the interaction, at stage 15, type II was present in the extracellular compartment basal to the epithelium. After completion of the interaction, collagen type II was no longer detectable at the interface even though it was readily detectable in the vitreous humor, cornea, and perinotochordal sheath, and subsequently will be expressed by the chondrogenic tissue itself as overt differentiation commences. These results suggest that collagen type II might be causally involved in this particular epitheliomesenchymal interaction. Examination of the spatial and temporal patterns of collagen type II expression elsewhere in the developing craniofacial complex revealed a hitherto unreported pattern of distribution. In addition to its predictable locations (i.e., cornea, vitreous, and perinotochordal sheath) it was found to be present at certain other sites, for example, at the basal surfaces of some neuroepithelia. These additional locations are all known to be sites of chondrogenesis-promoting tissue interactions which result in the formation of the elements of the cartilaginous neurocranium (e.g., otic vesicle). Furthermore this spatial distribution exhibits a changing temporal pattern in that it is detectable at the time that the interactions are known to be taking place, but subsequently is no longer detectable by the immunohistochemical means employed. This definable pattern of transient collagen type II expression, occurring at very early stages of craniofacial development, is interpreted as reflecting one level of morphogenetic specification of chondrocranial/skull form in the developing vertebrate head.

Keratoconus corneal architecture after riboflavin/ultraviolet A cross-linking: Ultrastructural studies

PubMed Central

Almubrad, Turki; Paladini, Iacopo; Mencucci, Rita

2013-01-01

Purpose Study to investigate the effects of collagen cross-linking on the ultrastructural organization of the corneal stroma in the human keratoconus cornea (KC). Methods Three normal, three keratoconus (KC1, KC2, KC3), and three cross-linked keratoconus (CXL1, CXL2, CXL3) corneas were analyzed. The KC corneas were treated with a riboflavin-ultraviolet A (UVA) treatment (CXL) method described by Wollensak et al. Penetrating keratoplasty (PKP) was performed 6 months after treatment. All samples were processed for electron microscopy. Results The riboflavin-UVA-treated CXL corneal stroma showed interlacing lamellae in the anterior stroma followed by well-organized parallel running lamellae. The lamellae contained uniformly distributed collagen fibrils (CFs) decorated with normal proteoglycans (PGs). The CF diameter and interfibrillar spacing in the CXL cornea were significantly increased compared to those in the KC cornea. The PG area in the CXL corneas were significantly smaller than the PGs in the KC cornea. The epithelium and Bowman’s layer were also normal. On rare occasions, a thick basement membrane and collagenous pannus were also observed. Conclusions Corneal cross-linking leads to modifications of the cornea stroma. The KC corneal structure showed a modification in the CF diameter, interfibrillar spacing, and PG area. This resulted in a more uniform distribution of collagen fibrils, a key feature for corneal transparency. PMID:23878503

The effect of actinoquinol with hyaluronic acid in eye drops on the optical properties and oxidative damage of the rabbit cornea irradiated with UVB rays.

PubMed

Čejka, Čestmír; Luyckx, Jacques; Ardan, Taras; Pláteník, Jan; Širc, Jakub; Michálek, Jiří; Čejková, Jitka

2010-01-01

Irradiation of the cornea with UVB rays leads to its oxidative damage, swelling and increased light absorption. We investigated changes in the corneal optics (evaluated by changes of corneal hydration and light absorption) and microscopical disturbances of corneas irradiated with UVB rays as influenced by eye drops containing actinoquinol with hyaluronic acid. Rabbit corneas were irradiated with a daily dose of 0.5 or 1.01 J cm(-2) of UVB rays (312 nm) for 4 days. During irradiation, the eye drops were applied on the right eye and buffered saline (or hyaluronic acid) on the left eye. On day 5 the rabbits were sacrificed and the corneas examined spectrophotometrically for light absorption. The corneal thickness (hydration) was measured using a pachymeter. Corneas of some other rabbits were examined immunohistochemically. After buffered saline treatment UVB rays evoked changes in the corneal optics and induced oxidative damage of the corneas. After actinoquinol-hyaluronic acid application, these changes were diminished. Hyaluronic acid alone was less effective. In conclusion, actinoquinol-hyaluronic acid eye drops decreased changes in corneal optics and suppressed oxidative damage in the UVB-irradiated cornea. However, the effective corneal protection by these eye drops was limited to the lower UVB dose. © 2010 The Authors. Journal Compilation. The American Society of Photobiology.