Journal of Virology 2014-12-01

Establishment and characterization of an air-liquid canine corneal organ culture model to study acute herpes keratitis.

Rebecca M Harman, Leen Bussche, Eric C Ledbetter, Gerlinde R Van de Walle

Index: J. Virol. 88(23) , 13669-77, (2014)

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Abstract

Despite the clinical importance of herpes simplex virus (HSV)-induced ocular disease, the underlying pathophysiology of the disease remains poorly understood, in part due to the lack of adequate virus-natural-host models in which to study the cellular and viral factors involved in acute corneal infection. We developed an air-liquid canine corneal organ culture model and evaluated its susceptibility to canine herpesvirus type 1 (CHV-1) in order to study ocular herpes in a physiologically relevant natural host model. Canine corneas were maintained in culture at an air-liquid interface for up to 25 days, and no degenerative changes were observed in the corneal epithelium during cultivation using histology for morphometric analyses, terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL) assays, and transmission electron microscopy (TEM). Next, canine corneas were inoculated with CHV-1 for 48 h, and at that time point postinfection, viral plaques could be visualized in the corneal epithelium and viral DNA copies were detected in both the infected corneas and culture supernatants. In addition, we found that canine corneas produced proinflammatory cytokines in response to CHV-1 infection similarly to what has been described for HSV-1. This emphasizes the value of our model as a virus-natural-host model to study ocular herpesvirus infections.This study is the first to describe the establishment of an air-liquid canine corneal organ culture model as a useful model to study ocular herpesvirus infections. The advantages of this physiologically relevant model include the fact that (i) it provides a system in which ocular herpes can be studied in a virus-natural-host setting and (ii) it reduces the number of experimental animals needed. In addition, this long-term explant culture model may also facilitate research in other fields where noninfectious and infectious ocular diseases of dogs and humans are being studied.Copyright © 2014, American Society for Microbiology. All Rights Reserved.