Standardization of corneal alkali burn methodology in rabbits

Abstract

Purpose : To standardize alkali burns to the cornea with regards to whether the corneal endothelium, epithelium, and stroma were injured, which has a critical role in the subsequent corneal wound healing fibrosis response

Methods : New Zealand white rabbits under general ketamine-xylazine anesthesia and topical tetracaine anesthesia had alkali burns in one eye. A 5 mm diameter circle of Whatman #1 filter paper wetted with 30 microliters of control vehicle balanced salt solution (BSS) or 0.25N,0.5N,0.75N, or 1N sodium hydroxide (NaOH) in BSS was applied for 15 seconds prior to irrigation in corneas.Corneas were evaluated with standardized slit lamp photographs and triplex immunohistochemistry for keratocytes (keratocan),mesenchymal cells (vimentin),and myofibroblasts (alpha-smooth muscle actin)

Results : When 0.5N NaOH was used, the injury, myofibroblast generation, and fibrosis were limited to the anterior stroma. 0.75N NaOH use also triggered injury to Descemet’s membrane and the corneal endothelium with fibrosis of the full-thickness stroma.1N NaOH use produced more significant stromal fibrosis and stimulated corneal neovascularization and persistent epithelial defects

Conclusions : Alkali burns are commonly used in corneal wound healing studies and investigators have used different methods that varied in sodium hydroxide concentration, application methods,and duration of exposure. A critical factor in the subsequent corneal healing response, including myofibroblast generation and fibrosis localization,was whether Descemet’s membrane and the endothelium were injured during the initial exposure. 0.5N NaOH exposures produced injuries confined to the epithelium and stroma, and anterior stromal myofibroblasts and fibrosis were produced. However, 0.75N or 1N NaOH also injured Descemet’s membrane and the endothelium,with myofibroblast generation and fibrosis noted in the full thickness of the cornea.DOD grant VR210001

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.

 

Immunohistochemistry for keratocan, vimentin,and a-SMA in corneas exposed to NaOH in BSS for 15 seconds at 1 month post-treatment. All corneas treated with 0.75N or 1N NaOH had damage to the endothelium and stromal fibrosis present in a spotty distribution from the anterior to posterior cornea. The central stroma in each cornea has less fibrosis than the anterior and posterior stroma, likely related to the bioavailability of activated TGF-b. Arrowheads in a-SMA: myofibroblasts,in vimentin panels: mesenchymal cells

 

Arrows indicate CNV.