Cornea - ARVO

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ARVO 2013 Annual Meeting Abstracts by Scientific Section/Group - CorneaEvaluation of transcorneal iontophoresis of riboflavin for cornealcollagen cross-linkingAlejandro Arboleda 1, 2 , Laura Kowalczuk 3 , Michèle Savoldelli 4 ,Christophe Klein 3 , Sophia Ladraa 3 , Jean-Marie A. Parel 1, 4 , FrancineF. Behar-Cohen 3, 4 . 1 Ophthalmic Biophysics Center, Dept. ofOphthalmology, Bascom Palmer Eye Institute, University of MiamiMiller School of Medicine, Miami, FL; 2 Dept. of BiomedicalEngineering, University of Miami College of Engineering, CoralGables, FL; 3 INSERM UMRS872: Physiopathology of OcularDiseases: Therapeutic Innovations, Centre de Recherche desCordeliers, Pierre et Marie Curie University, Paris DescartesUniversity, Paris, France; 4 AP-HP Hôtel-Dieu, Dept. ofOphthalmology, Sorbonne Paris Cité, Paris Descartes University,Faculty of Medicine, Paris, France.Purpose: To determine whether two riboflavin solutions can bedelivered into the cornea through iontophoresis prior to collagencross-linking. Two conditions, epithelium-on and epithelium-off,were compared to determine which allowed for the most diffusion ofriboflavin throughout the cornea.Methods: A Coulomb-controlled iontophoresis (CCI) generator anda corneal probe were used to deliver either 0.1% riboflavindextranT5000or 0.1% riboflavin-phosphate into the eyes of 8-weekoldLewis rats under either epithelium-on (epi-on) or epithelium-off(epi-off) conditions. Within 30 minutes of CCI (1.77 mA/cm 2 for 4minutes), corneas were flat-mounted for fluorescence imaging usinga confocal microscope (LSM 710, Zeiss). The distribution ofriboflavin in the cornea was evaluated by recording the fluorescenceintensity. Aqueous humors (Aq.H) were also collected to assayriboflavin concentration by fluorometry using a multilab counter(Wallac Victor 1420, Perkin Elmer).Results: Iontophoresis delivered both riboflavin solutions throughoutthe whole cornea for the epi-off condition. For the epi-on case, onlyriboflavin-phosphate was successfully delivered to the cornea (Figure1). Corneal imaging and Aq.H assay both demonstrated thatriboflavin-phosphate delivery is more efficient than riboflavindextrandelivery. The riboflavin concentrations in Aq.H were: epi-onriboflavin-phosphate, 0.92 µg/mL; epi-on riboflavin-dextran, 0.55µg/mL; epi-off riboflavin-phosphate, 13.6 µg/mL; and epi-offriboflavin-dextran, 4.73 µg/mL.Conclusions: Iontophoresis can be used to efficiently deliverriboflavin into the cornea. The epi-off condition enhances diffusionof riboflavin into the stroma and aqueous humor. Transcornealdelivery is only possible with riboflavin-phosphate. The efficiency ofCXL through an intact epithelium, which absorbs most UV radiation,remains to be demonstrated.Figure 1: White shows riboflavin fluorescence: the whiter, the higherthe riboflavin concentration (A) Riboflavin-dextran diffusion in thecornea for the epi-off condition (B) Riboflavin-dextran diffusion inthe cornea for the epi-on condition (C) Riboflavin-phosphatediffusion in the cornea for the epi-off condition (D) Riboflavinphosphatediffusion in the cornea for the epi-on conditionCommercial Relationships: Alejandro Arboleda, None; LauraKowalczuk, None; Michèle Savoldelli, None; Christophe Klein,None; Sophia Ladraa, None; Jean-Marie A. Parel, CROMA (F),InnFocus (F), Abeamed (F), University of Miami (P); Francine F.Behar-Cohen, Inserm/Univesrité ParisDescartes (P)Support: Agence Nationale de la Recherche Grant ANR-11-RPIB-020, Florida Lions Eye Bank, NIH Center Grant P30EY14801,Research to Prevent Blindness, Florida-Georgia Louis StokesAlliance for Minority Participation, Henri and Flore LesieurFoundation (JMP)Program Number: 4073Presentation Time: 9:30 AM - 9:45 AMCorneal biomechanical properties in corneal collagen crosslinking(CXL) at high fluencesArthur Hammer 1 , Olivier Richoz 1 , David Tabibian 1 , FlorenceHoogewoud 1 , Farhad Hafezi 1, 2 . 1 Ophthalmology, Geneva UniversityHospitals, Geneva, Switzerland; 2 Ophthalmology, Doheny EyeInstitute, Keck school of medicine USC, Los Angeles, CA.Purpose: The current intensity/irradiation profile for collagen crosslinking(3 mW/cm2 for 30 minutes) has been in clinical use since1999 in a multitude of studies. Lately, different irradiation profileshave emerged, with the intention to lower irradiation time whilemaintaining the total energy dose (Bunsen-Roscoe law). Little isknown whether these modified irradiation profiles will lead to thesame increase in biomechanical stiffness observed in the originalprotocol.Methods: We investigated the biomechanical properties in ex vivoporcine corneas at 3 (n = 10), 9 (n = 10), and 18 (N = 12) mW/cm2for various time periods (30, 10, 5 minutes), while maintaining thetotal energy dose identical (Bunsen-Roscoe law). Prior to irradiation,riboflavin 0.1% was applied on the de-epithelialized cornea for 20minutes. Controls (n = 11) were treated similarly, but without UV-Airradiation. Stress-strain measurements were performed using anextensometer (Zwick Roell, Model ZO.05, Zwick GmbH & Co. KG,Ulm, Germany).Results: We observed a decrease in Young’s modulus withincreasing UV-A intensity (fluence). Young’s modulus at 4 %, 6 %, 8% and 10 % strain at 3 mW/cm2 was 1.66 N, 0.75 N, 0.44 N and 0.29N respectively. At 9 mW/cm2, we measured 1.52 N, 0.7 N, 0.39 Nand 0.24 N, whereas at 18 mW/cm2, we detected 1.2 N, 0.57 N, 0.33N and 0.21 N for 18 mW. Controls showed 0.72 N, 0.37 N, 0.37 Nand 0.16 N.Conclusions: The increase in biomechanical strength of the porcinecornea following CXL diminishes with increasing fluence/decreasingirradiation time, even if the total energy dose is maintained.The generation of chemical bonds during cross-linking is an oxygendependentprocess that depends on intra-stromal oxygenconcentration. Oxygen diffusion capacity is limited within the stromaand the increased oxygen consumption at higher fluence/lowerirradiation time might become a limiting factor for cross-linking,leading to the observed decrease in treatment efficacy.Commercial Relationships: Arthur Hammer, None; OlivierRichoz, None; David Tabibian, None; Florence Hoogewoud, None;Farhad Hafezi, Schwind (F), Ziemer (F), PCT/CH 2012/000090 (P)Program Number: 4074©2013, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permissionto reproduce any abstract, contact the ARVO Office at arvo@arvo.org.
ARVO 2013 Annual Meeting Abstracts by Scientific Section/Group - CorneaPresentation Time: 9:45 AM - 10:00 AMSoluble collagen protects the corneal fibrils during riboflavincrosslinkingMarcos Garza-Madrid 1, 2 , Jennifer Elisseeff 1 . 1 Translational TissueEngineering Center, Wilmer Eye Institute, Johns Hopkins School ofMedicine, Baltimore, MD; 2 Ophthalmology and Visual SciencesResearch Chair, School of Medicine and Health Sciences,Tecnológico de Monterrey, Monterrey, Mexico.Purpose: Cornea crosslinking with riboflavin and UV-A is a novelprocedure able to stop the progression of keratoconus. Our aim is tocharacterize and prevent the fibrillar damage associated with thisprocedure.Methods: Fresh bovine corneas were obtained from a localslaughterhouse. Six mm biopsies, devoid of epithelium andendothelium were treated with a Riboflavin isotonic solution with orwithout soluble collagen. The tissue was then crosslinked using UV-A light. Hydration was maintained by adding crosslinking solutionperiodically. Samples were then either processed for TEM oranalyzed through DSC to identify the temperature at which themolecular structure breaks down. Fibril counting was performedmanually on a minimum of 180 fibrils per group.Results: After 30 minutes of crosslinking the denaturationtemperature of the corneas increased from 66.77±0.92°C to70.94±1.72°C (p
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