New Tools for the Evaluation of Rat Ocular Surface Changes

Abstract

Purpose: : To develop a short–time model of ocular toxicity in order to assess the usefulness of a set of ocular surface evaluating methods

Methods: : Two groups of 3 rats each received benzalkonium chloride (BAC) at the classical 0.01% eye–drop concentration and at the supratoxic 0.5% dose in the right eye during 11 days. At days 0, 3, 8, 11, 15, 18, 20, 22, 31, 38, 45 and 52, the cotton thread method and the fluorescein test were used so as to monitor the rat lachrymal secretion and evaluate corneal damages, respectively; the new generation in vivo confocal microscopy – HRT–II cornea module – was used to follow–up corneal lesions, changes in cellular density, and neovascularization on treated and control corneas. MUC–5AC expression was analyzed on conjunctival impression cytology specimens and conjunctival imprints were also processed by flow cytometry for cell cycle analysis.

Results: : 0.5% BAC induced epithelial abrasion, alteration of the endothelium and the stroma, stromal cell infiltration and decreased conjunctival MUC–5AC expression from D3 to D11. It also induced persistent corneal neovascularization from D8 to D52. 0.01% BAC induced apoptosis as detected on conjunctival imprints.

Conclusions: : New generation in vivo confocal microscopy provides in a non–invasive way high–resolution in–situ imaging of ocular structures, especially inflammatory cell infiltration and blood vessel growth. We validated the usefulness of impression cytology to follow ocular surface changes in rat using confocal microscopy and flow cytometry. Such a set of combined technologies could also serve as a basis to supplement the classical Draize test in toxicological studies.

Keywords: ocular irritancy/toxicity testing • imaging methods (CT, FA, ICG, MRI, OCT, RTA, SLO, ultrasound) • neovascularization