Development Of An Experimental Ex-Vivo Model Of Corneal Opacity And Wound Healing
Published 2025 - 43rd Congress of the ESCRS
Reference: PP20.10 | Type: Free paper | DOI: 10.82333/ze5d-qx72
Authors: Jacek P. Szaflik* 1 , Karolina Pielak-Modrzejewska 2 , Emilia Babula 2 , Katarzyna Nowik 2 , Agata J. Ordon 1 , Tomasz Gałecki 2 , Justyna Izdebska 1 , Anna Kamińska 1
1Department of Ophthalmology,Medical University of Warsaw,Warsaw,Poland;SPKSO Ophthalmic University Hospital ,Warsaw,Poland;Laser Eye Microsurgery Centre Clinic of Prof. Jerzy Szaflik,Warsaw,Poland, 2Department of Ophthalmology,Medical University of Warsaw,Warsaw,Poland;SPKSO Ophthalmic University Hospital ,Warsaw,Poland
Purpose
To develop an experimental ex-vivo model of corneal stromal scarring able to reproduce corneal opacity as seen in corneal haze formation. To perform histopathologic tissue analysis and characterize the complex cascade of corneal stromal opacity.
Setting
Institute of Applied Ophthalmobiology (IOBA), Universitiy of Valladolid, Valladolid, Spain
Methods
Enuclated domestic pig eyes obtained from local slaughterhouse and preserved in cell culture medium. To produce corneal opacities air-assisted deep manual keratectomy DMAK, DMAK+manual scraping MS, and DMAK+TGFβ1 exposure performed in two eye groups with/without epithelial wound. Corneal tissues maintained in culture for 7days at 37°C, 5%CO2, and 95%humidity. Epithelial wound closure evaluated daily with 2% fluorescein, images analyzed with ImageJ® software.Corneal opacity assessed with a nominal visual scale and weighted Kappa-Cohen coefficient. Histopathological stainings and immunofluorescence against myofibroblast marker α-SMA performed in tissue sections, results under different conditions compared. 3 independent experiments conducted
Results
The ex-vivo model was successfully maintained in culture for 7 days. Delayed epithelial wound closure in mm2 was observed in eyes treated with TGFβ1 compared with controls (p<0.05 t Student). Higher opacity scale scores were noted in eyes where TGFβ1 was applied and MS + DMAK were performed. A weighted Kappa-Cohen coefficient of 0,68 (CI 95% 0,36-0,99) and an 80,56% inter-observer agreement were obtained after the opacity scale applied to grade opacity. Histopathological analysis demonstrated disorganization of collagen fibers in the corneal tissue. Immunofluorescence showed reactivity with α-SMA antibody as a myofibroblast marker in the eyes where DMAK was performed, being greater in the eyes with TGFβ1
Conclusions
It’s possible to develop an experimental ex-vivo model of corneal stromal scarring and opacity using porcine eyes that represents features of stromal wound healing that leads to corneal haze formation. Further studies are needed to evaluate the applicability of this model in order to consider future preventive and therapeutic approaches for corneal opacities and haze.