Transcorneal Freezing in Aged Rabbits as a Platform for Evaluating Corneal Endothelial Cell Therapeutics.

Purpose: Transcorneal freezing is a common technique used in rabbits to induce damage to the corneal endothelium. Previous studies have been performed with a range of freezing temperatures, times, and rabbit ages. Here, we aimed to characterize the aged rabbit endothelium after transcorneal freezing to establish an innate corneal endothelial cell regrowth rate and propose it as a mechanism for evaluation of therapeutic efficacy in rabbit models.

Methods: Central corneas of anesthetized New Zealand White rabbits (n=3) aged 18-24 months were exposed to nitrous oxide cooled probes for 30 seconds. Animals were assessed by in vivo confocal microscopy, applanation tonometry, specular microscopy, optical coherence tomography, and histology. The contralateral eye acted as a control. Images were taken immediately before and after injury and on days 2, 4, 7, 11, and 14.

Results: Following transcorneal freezing, there was a significant decrease in corneal endothelium density and a temporary increase in corneal thickness. Endothelial density decreased by 95% immediately after injury compared to controls and showed linear recovery over 14 days, reaching a 38% reduction by day 14. There was a significant increase in pleomorphism across all time points post-injury. Conversely, corneal thickness increased two days post injury but recovered at all later time points. Intraocular pressure was not affected throughout.

Conclusions: This corneal endothelium injury platform is ideal for injury and therapeutic research as it can be rapidly performed, and has minimal impact on corneal thickness and intraocular pressure. Due to innate rabbit endothelial regrowth, it is vital to establish corneal endothelial recovery rate before evaluating therapeutics for efficacy in this model system.