Application of Collagen I and IV in Bioengineering Transparent Ocular Tissues
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Generation and Characterization of a Novel Mouse Line, Keratocan-Rtta (Kerart), for Corneal Stroma and Tendon Research
Cornea Generation and Characterization of a Novel Mouse Line, Keratocan-rtTA (KeraRT), for Corneal Stroma and Tendon Research Yujin Zhang,1 Winston W.-Y. Kao,2 Yasuhito Hayashi,3 Lingling Zhang,1 Mindy Call,2 Fei Dong,2 Yong Yuan,2 Jianhua Zhang,2 Yen-Chiao Wang,1 Okada Yuka,1,4 Atsushi Shiraishi,3 and Chia-Yang Liu1 1School of Optometry, Indiana University, Bloomington, Indiana, United States 2Edith J. Crawley Vision Research Center/Department of Ophthalmology, University of Cincinnati College of Medicine, Cincinnati, Ohio, United States 3Department of Ophthalmology, School of Medicine, Ehime University, Ehime, Japan 4Department of Ophthalmology, School of Medicine, Wakayama Medical University, Wakayama, Japan RT Correspondence: Yujin Zhang, Indi- PURPOSE. We created a novel inducible mouse line Keratocan-rtTA (Kera ) that allows ana University School of Optometry, specific genetic modification in corneal keratocytes and tenocytes during development and in 800 Atwater Avenue, Bloomington, adults. IN 47405, USA; [email protected]. METHODS. A gene-targeting vector (Kera- IRES2-rtTA3) was constructed and inserted right after Chia-Yang Liu, Indiana University the termination codon of the mouse Kera allele via gene targeting techniques. The resulting RT RT School of Optometry, 800 Atwater Kera mouse was crossed to tet-O-Hist1H2B-EGFP (TH2B-EGFP) to obtain Kera /TH2B-EGFP Avenue, Bloomington, IN 47405, compound transgenic mice, in which cells expressing Kera are labeled with green USA; fluorescence protein (GFP) by doxycycline (Dox) induction. The expression patterns of [email protected]. RT RT GFP and endogenous Kera were examined in Kera /TH2B-EGFP. Moreover, Kera was bred Submitted: July 21, 2017 with tet-O-TGF-a to generate a double transgenic mouse, KeraRT/tet-O-TGF-a, to overexpress Accepted: August 16, 2017 TGF-a in corneal keratocytes upon Dox induction. -
Cultivating a Cure for Blindness
news and views were taken in a l-mm2 biopsy from the good eye, then they were cultured and, on second Cultivating a cure passage, they formed a tightly packed and communicating (confluent) monolayer of cells. For grafting, the conjunctiva! epi for blindness thelium was completely removed from the cornea and limbus of the recipient eye, and replaced with a slightly larger monolayer of Stuart Hodson cultured limbal epithelium. The eyes were Damaged corneas can often be repaired using donor grafts, but if the then covered with therapeutic soft contact damage is too great the graft wlll be rejected. This may change with the lenses, and tightly patched for several days. development of a method to Inhibit rejection which uses cultivated cells. The results after the limbal epithelial graft were very promising. The grafted epithelium he human cornea has a special proper epithelium can mould a smooth apical and was stable, transparent, multi-layered and Tty known as immune privilege, which basal surface, the limbal epithelial stem cells smooth. One of the patients had suffered an allows tissue grafts from donors to be do not form such a smooth surface. alkali burn to his left eye ten years earlier, and carried out without the usual problems of If the corneal epithelium is lost it can be had undergone three previous unsuccessful immune rejection. However, immune privi functionally regenerated by the limbal stem corneal grafts. Before the treatment he had lege is lost at the perimeter of the cornea - cells. But if both the corneal and limbal continual severe corneal vascularization the limb us of the eye - where the transpar epithelia are lost, the corneal surface is re ( development of blood vessels) and persis ent corneal stroma meets the opaque sclera colonized by the other neighbour of the tent ulceration, and the eye was painful and (Fig. -
Quantitative Assessment of Central and Limbal Epithelium After Long
Eye (2016) 30, 979–986 © 2016 Macmillan Publishers Limited All rights reserved 0950-222X/16 www.nature.com/eye 1,5 1,5 1 Quantitative RK Prakasam , BS Kowtharapu , K Falke , CLINICAL STUDY K Winter2,3, D Diedrich4, A Glass4, A Jünemann1, assessment of central RF Guthoff1 and O Stachs1 and limbal epithelium after long-term wear of soft contact lenses and in patients with dry eyes: a pilot study Abstract Purpose Analysis of microstructural Eye (2016) 30, 979–986; doi:10.1038/eye.2016.58; alterations of corneal and limbal epithelial published online 22 April 2016 cells in healthy human corneas and in other ocular conditions. Introduction Patients and methods Unilateral eyes of three groups of subjects include healthy The X, Y, Z hypothesis1 explains cell mechanism volunteers (G1, n = 5), contact lens wearers that is essential for the renewal and maintenance 1Department of (G2, n = 5), and patients with dry eyes of the corneal epithelium. This hypothesis Ophthalmology, University = proposes that the loss of corneal epithelial of Rostock, Rostock, (G3, n 5) were studied. Imaging of basal Germany (BC) and intermediate (IC) epithelial cells surface cells (Z) can be maintained by the from central cornea (CC), corneal limbus proliferation of basal epithelial cells (X), and the 2Faculty of Medicine, centripetal movements of the peripheral (CL) and scleral limbus (SL) was obtained by Institute of Anatomy, epithelial cells (Y). By utilizing this mechanism, University of Leipzig, in vivo confocal microscopy (IVCM). An it is also possible to categorize both disease and Leipzig, Germany appropriate image analysis algorithm was therapies according to the specific component 3 used to quantify morphometric parameters involved.1 Therefore it is vital to understand the Institute for Medical including mean cell area, compactness, Informatics, Statistics and cellular structures of both central and limbal Epidemiology (IMISE), solidity, major and minor diameter, and epithelial cells in normal and in various corneal University of Leipzig, maximum boundary distance. -
Development of in Vitro Corneal Models: Opportunity for Pharmacological Testing
Review Development of In Vitro Corneal Models: Opportunity for Pharmacological Testing Valentina Citi 1, Eugenia Piragine 1, Simone Brogi 1,* , Sara Ottino 2 and Vincenzo Calderone 1 1 Department of Pharmacy, University of Pisa, Via Bonanno 6, 56126 Pisa, Italy; [email protected] (V.C.); [email protected] (E.P.); [email protected] (V.C.) 2 Farmigea S.p.A., Via G.B. Oliva 6/8, 56121 Pisa, Italy; [email protected] * Correspondence: [email protected]; Tel.: +39-050-2219-613 Received: 24 October 2020; Accepted: 30 October 2020; Published: 2 November 2020 Abstract: The human eye is a specialized organ with a complex anatomy and physiology, because it is characterized by different cell types with specific physiological functions. Given the complexity of the eye, ocular tissues are finely organized and orchestrated. In the last few years, many in vitro models have been developed in order to meet the 3Rs principle (Replacement, Reduction and Refinement) for eye toxicity testing. This procedure is highly necessary to ensure that the risks associated with ophthalmic products meet appropriate safety criteria. In vitro preclinical testing is now a well-established practice of significant importance for evaluating the efficacy and safety of cosmetic, pharmaceutical, and nutraceutical products. Along with in vitro testing, also computational procedures, herein described, for evaluating the pharmacological profile of potential ocular drug candidates including their toxicity, are in rapid expansion. In this review, the ocular cell types and functionality are described, providing an overview about the scientific challenge for the development of three-dimensional (3D) in vitro models. -
Vocabulario De Morfoloxía, Anatomía E Citoloxía Veterinaria
Vocabulario de Morfoloxía, anatomía e citoloxía veterinaria (galego-español-inglés) Servizo de Normalización Lingüística Universidade de Santiago de Compostela COLECCIÓN VOCABULARIOS TEMÁTICOS N.º 4 SERVIZO DE NORMALIZACIÓN LINGÜÍSTICA Vocabulario de Morfoloxía, anatomía e citoloxía veterinaria (galego-español-inglés) 2008 UNIVERSIDADE DE SANTIAGO DE COMPOSTELA VOCABULARIO de morfoloxía, anatomía e citoloxía veterinaria : (galego-español- inglés) / coordinador Xusto A. Rodríguez Río, Servizo de Normalización Lingüística ; autores Matilde Lombardero Fernández ... [et al.]. – Santiago de Compostela : Universidade de Santiago de Compostela, Servizo de Publicacións e Intercambio Científico, 2008. – 369 p. ; 21 cm. – (Vocabularios temáticos ; 4). - D.L. C 2458-2008. – ISBN 978-84-9887-018-3 1.Medicina �������������������������������������������������������������������������veterinaria-Diccionarios�������������������������������������������������. 2.Galego (Lingua)-Glosarios, vocabularios, etc. políglotas. I.Lombardero Fernández, Matilde. II.Rodríguez Rio, Xusto A. coord. III. Universidade de Santiago de Compostela. Servizo de Normalización Lingüística, coord. IV.Universidade de Santiago de Compostela. Servizo de Publicacións e Intercambio Científico, ed. V.Serie. 591.4(038)=699=60=20 Coordinador Xusto A. Rodríguez Río (Área de Terminoloxía. Servizo de Normalización Lingüística. Universidade de Santiago de Compostela) Autoras/res Matilde Lombardero Fernández (doutora en Veterinaria e profesora do Departamento de Anatomía e Produción Animal. -
Comparative Analysis of a Teleost Skeleton Transcriptome Provides Insight Into Its Regulation
Accepted Manuscript Comparative analysis of a teleost skeleton transcriptome provides insight into its regulation Florbela A. Vieira, M.A.S. Thorne, K. Stueber, M. Darias, R. Reinhardt, M.S. Clark, E. Gisbert, D.M. Power PII: S0016-6480(13)00264-5 DOI: http://dx.doi.org/10.1016/j.ygcen.2013.05.025 Reference: YGCEN 11541 To appear in: General and Comparative Endocrinology Please cite this article as: Vieira, F.A., Thorne, M.A.S., Stueber, K., Darias, M., Reinhardt, R., Clark, M.S., Gisbert, E., Power, D.M., Comparative analysis of a teleost skeleton transcriptome provides insight into its regulation, General and Comparative Endocrinology (2013), doi: http://dx.doi.org/10.1016/j.ygcen.2013.05.025 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. 1 Comparative analysis of a teleost skeleton transcriptome 2 provides insight into its regulation 3 4 Florbela A. Vieira1§, M. A. S. Thorne2, K. Stueber3, M. Darias4,5, R. Reinhardt3, M. 5 S. Clark2, E. Gisbert4 and D. M. Power1 6 7 1Center of Marine Sciences, Universidade do Algarve, Faro, Portugal. 8 2British Antarctic Survey – Natural Environment Research Council, High Cross, 9 Madingley Road, Cambridge, CB3 0ET, UK. -
Supplementary Table 1: Adhesion Genes Data Set
Supplementary Table 1: Adhesion genes data set PROBE Entrez Gene ID Celera Gene ID Gene_Symbol Gene_Name 160832 1 hCG201364.3 A1BG alpha-1-B glycoprotein 223658 1 hCG201364.3 A1BG alpha-1-B glycoprotein 212988 102 hCG40040.3 ADAM10 ADAM metallopeptidase domain 10 133411 4185 hCG28232.2 ADAM11 ADAM metallopeptidase domain 11 110695 8038 hCG40937.4 ADAM12 ADAM metallopeptidase domain 12 (meltrin alpha) 195222 8038 hCG40937.4 ADAM12 ADAM metallopeptidase domain 12 (meltrin alpha) 165344 8751 hCG20021.3 ADAM15 ADAM metallopeptidase domain 15 (metargidin) 189065 6868 null ADAM17 ADAM metallopeptidase domain 17 (tumor necrosis factor, alpha, converting enzyme) 108119 8728 hCG15398.4 ADAM19 ADAM metallopeptidase domain 19 (meltrin beta) 117763 8748 hCG20675.3 ADAM20 ADAM metallopeptidase domain 20 126448 8747 hCG1785634.2 ADAM21 ADAM metallopeptidase domain 21 208981 8747 hCG1785634.2|hCG2042897 ADAM21 ADAM metallopeptidase domain 21 180903 53616 hCG17212.4 ADAM22 ADAM metallopeptidase domain 22 177272 8745 hCG1811623.1 ADAM23 ADAM metallopeptidase domain 23 102384 10863 hCG1818505.1 ADAM28 ADAM metallopeptidase domain 28 119968 11086 hCG1786734.2 ADAM29 ADAM metallopeptidase domain 29 205542 11085 hCG1997196.1 ADAM30 ADAM metallopeptidase domain 30 148417 80332 hCG39255.4 ADAM33 ADAM metallopeptidase domain 33 140492 8756 hCG1789002.2 ADAM7 ADAM metallopeptidase domain 7 122603 101 hCG1816947.1 ADAM8 ADAM metallopeptidase domain 8 183965 8754 hCG1996391 ADAM9 ADAM metallopeptidase domain 9 (meltrin gamma) 129974 27299 hCG15447.3 ADAMDEC1 ADAM-like, -
A Technique for Obtaining Basal Corneal Epithelial Cells
Reports A Technique for Obtaining Basal Corneal Epithelial Cells V. Trinkaus-Randall and I. K. Gipson A technique has been developed for obtaining a cell suspen- basal cells can be freed from the basal laminae after sion enriched (89%) in basal corneal epithelial cells. Eleven an incubation with Dispase II.8 millimeter corneal buttons were removed and placed in Materials and Methods. All investigations involving culture medium containing low (10 nM) calcium. The animals reported in this study conform to the ARVO posterior half of the stroma was removed with forceps. Resolution on the Use of Animals in Research. New Three superficial cuts were made with a Bard-Parker blade on the anterior half of the cornea, which was then incubated Zealand white rabbits were killed by administering 5 for 18 hr at 35°C. Nonadherent cells were brushed off after ml pentobarbital (325 mg) intravenously. An outline the incubation and basal cells were harvested after a 1-hr was made on the cornea with an 11-mm trephine incubation in Dispase II. Cell viability estimated by Eryth- and corneal buttons were cut free with scissors. The rocin /? exclusion was 90%. Further evidence of viability posterior half of the stroma then was pulled away was that the cells adhered to their native substrate, the from the cornea with jewelers' forceps. Three evenly denuded basal lamina. The authors protocol provides a spaced, superficial cuts (1 mm in length) were made method for analyzing the biochemistry of a known population on the cornea with a small scalpel to facilitate the of epithelial cells and makes available a defined source of penetration of the medium. -
Maintaining the Cornea and the General Physiological Environment in Visual Neurophysiology Experiments
Journal of Neuroscience Methods 109 (2001) 153–166 www.elsevier.com/locate/jneumeth Maintaining the cornea and the general physiological environment in visual neurophysiology experiments Andrew B. Metha a, Alison M. Crane b , H. Grady Rylander III c, Sharon L. Thomsen d, Duane G. Albrecht b,* a Department of Optometry and Vision Sciences, The Uni6ersity of Melbourne, Carlton, Victoria 3053, Australia b Department of Psychology, Uni6ersity of Texas, Austin, TX 78712, USA c Department of Electrical and Computer Engineering, Uni6ersity of Texas, Austin, TX 78712, USA d Biomedical Engineering Program, Uni6ersity of Texas, Austin, TX 78712, USA Received 30 January 2001; received in revised form 4 June 2001; accepted 4 June 2001 Abstract Neurophysiologists have been investigating the responses of neurons in the visual system for the past half-century using monkeys and cats that are anesthetized and paralyzed, with the non-blinking eyelids open for prolonged periods of time. Impermeable plastic contact lenses have been used to prevent dehydration of the corneal epithelium, which would otherwise occur in minutes. Unfortunately, such lenses rapidly introduce a variety of abnormal states that lead to clouding of the cornea, degradation of the retinal image, and premature termination of the experiment. To extend the viability of such preparations, a new protocol for maintenance of corneal health has been developed. The protocol uses rigid gas permeable contact lenses designed to maximize gas transmission, rigorous sterile methods, and a variety of methods for sustaining and monitoring the overall physiology of the animal. The effectiveness of the protocol was evaluated clinically by ophthalmoscopy before, during, and after the experiments, which lasted 8–10 days. -
3D Functional Corneal Stromal Tissue Equivalent Based on Corneal Stromal Stem Cells and Multi-Layered Silk Film Architecture
RESEARCH ARTICLE 3D Functional Corneal Stromal Tissue Equivalent Based on Corneal Stromal Stem Cells and Multi-Layered Silk Film Architecture Chiara E. Ghezzi1, Benedetto Marelli1, Fiorenzo G. Omenetto1, James L. Funderburgh2, David L. Kaplan1* 1 Department of Biomedical Engineering, Tufts University, Medford, Massachusetts, United States of America, 2 Department of Ophthalmology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, United States of America * [email protected] a1111111111 a1111111111 a1111111111 Abstract a1111111111 a1111111111 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 OPEN ACCESS (hCSSCs), would generate 3D functional corneal stroma tissue equivalents, in comparison Citation: Ghezzi CE, Marelli B, Omenetto FG, to previously developed 2D approaches. Silk film contact guidance was used to control the Funderburgh JL, Kaplan DL (2017) 3D Functional alignment and distribution of hCSSCs on RGD-treated single porous silk films, which were Corneal Stromal Tissue Equivalent Based on then stacked in an orthogonally, multi-layered architecture and cultured for 9 weeks. These Corneal Stromal Stem Cells and Multi-Layered Silk Film Architecture. PLoS ONE 12(1): e0169504. systems were compared similar systems generated with human corneal fibroblasts (hCFs). doi:10.1371/journal.pone.0169504 Both cell types were viable and preferentially aligned along the biomaterial patterns for up to Editor: Dimitrios Karamichos, Oklahoma State 9 weeks in culture. H&E histological sections showed that the systems seeded with the University Center for Health Sciences, UNITED hCSSCs displayed ECM production throughout the entire thickness of the constructs. -
Changes in Epithelial and Stromal Corneal Stiffness Occur with Age
bioengineering Article Changes in Epithelial and Stromal Corneal Stiffness Occur with Age and Obesity Peiluo Xu 1, Anne Londregan 2 , Celeste Rich 3 and Vickery Trinkaus-Randall 3,4,* 1 Department of Biomedical Engineering, Boston University, Boston, MA 02115, USA; [email protected] 2 Department of Biochemistry/Molecular Biology, Boston University, Boston, MA 02215, USA; [email protected] 3 Department of Biochemistry, Boston University School of Medicine, Boston, MA 02115, USA; [email protected] 4 Department of Ophthalmology, Boston University School of Medicine, Boston, MA 02115, USA * Correspondence: [email protected] Received: 14 January 2020; Accepted: 4 February 2020; Published: 7 February 2020 Abstract: The cornea is avascular, which makes it an excellent model to study matrix protein expression and tissue stiffness. The corneal epithelium adheres to the basement zone and the underlying stroma is composed of keratocytes and an extensive matrix of collagen and proteoglycans. Our goal was to examine changes in corneas of 8- and 15-week mice and compare them to 15-week pre-Type 2 diabetic obese mouse. Nanoindentation was performed on corneal epithelium in situ and then the epithelium was abraded, and the procedure repeated on the basement membrane and stroma. Confocal imaging was performed to examine the localization of proteins. Stiffness was found to be age and obesity dependent. Young’s modulus was greater in the epithelium from 15-week mice compared to 8-week mice. At 15 weeks, the epithelium of the control was significantly greater than that of the obese mice. There was a difference in the localization of Crb3 and PKCζ in the apical epithelium and a lack of lamellipodial extensions in the obese mouse. -
Corneal Erosion?
What Is the Cornea? The cornea is the clear front window of the eye. It covers the iris (colored portion of the eye) and the round pupil, much like a watch crystal covers the face of a watch. The cornea is composed of five layers. The outermost surface layer is called the epithelium. Normal Eye Anatomy What Is a Corneal Abrasion? A corneal abrasion is an injury (a scratch, scrape or cut) to the corneal epithelium. Abrasions are commonly caused by fingernail scratches, paper cuts, makeup brushes, scrapes from tree or bush limbs, and rubbing of the eye. Some eye conditions, such as dry eye, increase the chance of an abrasion. You may experience the following symptoms with corneal abrasion: • Feeling of having something in your eye • Pain and soreness of the eye • Redness of the eye • Sensitivity to light • Tearing • Blurred vision To detect an abrasion on the cornea, your ophthalmologist (Eye M.D.) will use a special dye called fluorescein (pronounced FLOR-uh-seen) to illuminate the injury. How Is a Corneal Abrasion Treated? Treatment may include the following: • Patching the injured eye to prevent eyelid blinking from irritating the injury. • Applying lubricating eyedrops or ointment to the eye to form a soothing layer between the eyelid and the abrasion. • Using antibiotics to prevent infection. • Dilating (widening) the pupil to relieve pain. • Wearing a special contact lens to help healing. Minor abrasions usually heal within a day or two; larger abrasions usually take about a week. It is important not to rub the eye while it is healing.