Ex vivo excimer laser ablation of cornea guttata and ROCK inhibitor-aided endothelial recolonization of ablated central cornea.

Stefan Kassumeh,Gregory Moloney,Martin Dirisamer,Annabel Von Studnitz,Nikolaus Luft,Andreas Ohlmann,Rudolf Fuchshofer,Siegfried G Priglinger

doi:10.1111/aos.14366

Abstract

To determine whether excimer laser ablation of guttae is a viable strategy for removal of diseased tissue in Fuchs' endothelial corneal dystrophy (FECD) on excised human Descemet membranes and whether an excimer laser-created wound on healthy human corneas ex vivo is recolonized with corneal endothelial cells. Descemet membranes of FECD patients and corneal endothelium of normal human corneas were ablated ex vivo using an excimer laser licensed for glaucoma surgery. Specimens were kept in cell culture medium supplemented with 10μm of rho-kinase inhibitor ripasudil. Corneal endothelial cell regeneration was observed using light and electron scanning microscopy. Furthermore, the whole corneal samples were evaluated by haematoxylin/eosin staining and immunohistochemical analysis using antibodies against Na+ /K+ -ATPase. Guttae and corneal endothelium could be ablated with an excimer laser without total ultrastructural damage to theDescemet membrane or stroma. Nearly complete endothelial wound closure was accomplished after 26-38days in treated corneas. Light and electron scanning microscopy suggested the establishment of a layer of flat endothelial cells. Additionally, Na+ /K+ -ATPase expression could only be observed on the inner side of the Descemet membrane. Our proof of concept study demonstrated that excimer lasers can be used to ablate diseased tissue from excised FECD Descemet membranes ex vivo. Additionally, corneal endothelial cells recolonize a previously ablated endothelial area in healthy human corneas ex vivo under treatment with ripasudil. Thus, our results are the first experimental basis to further investigate the feasibility of an excimer laser ablation as a graftless FECD treatment option.

Highlights

Fuchs’ endothelial corneal dystrophy (FECD) is the most common dystrophy of the corneal endothelium leading to a significant decline in visual acuity
Keeping them in cell culture medium supplemented with 10 μM of ripasudil for 2 days, migration of endothelial cells from the edges of the descemetorhexis into the wounded area was observed by electron microscopy (Fig. 1A,B)
Laser ablation of guttae and Descemet membranes additional posterior banded layer To analyse whether the pulsed excimer laser beam can ablate guttae and the additional posterior banded layer of the Descemet membrane of patients suffering from FECD, preparations of patients undergoing Descemet membrane endothelial keratoplasty were whole mounted on a cell culture dish and stained with trypan blue prior to ex vivo treatment

Summary

Introduction

Fuchs’ endothelial corneal dystrophy (FECD) is the most common dystrophy of the corneal endothelium leading to a significant decline in visual acuity. As Fuchs’ endothelial corneal dystrophy develops, the characteristic proteinaceous lesions known as guttae accumulate as excrescences on the Descemet membrane (Abbott et al 1981). In early stages, they may produce light scatter and visual blur, when confluent and enlarged, they are not compatible with endothelial cell homeostasis and lead to cell death. One hypothesis suggests unknown processes may lead to a transformation of endothelial cells to a fibroblast-like phenotype, secreting collagen fibrils and forming the PBL Focal accelerations of those transformation processes are suggested to lead to guttae (Iwamoto & DeVoe 1971; Adamis et al 1993)

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