Evaluation of the Algerbrush II rotating burr as a tool for inducing limbal stem cell deficiency

Abstract

Purpose : Animal models of limbal stem cell deficiency (LSCD) typically involve chemical methods of epithelial debridement performed in conjunction with a limbectomy. Given variable results, we have presently evaluated a mechanical debridement technique based upon use of the Algerbrush II rotating burr.

Methods : Epithelial debridement was initially studied using cadaveric eyes of New Zealand White rabbits in situ (4 eyes for each method). Two different Algerbrush methods were tested; application of a 1 mm burr to the limbus, followed by application of a 2.5 mm burr to the cornea, or, application of the 2.5 mm burr to the entire ocular surface. Debrided eyes were immediately fixed and processed for histology (H&E staining). Controls consisted of non-wounded tissue and limbectomized tissue treated with 100% (v/v) n-heptanol. Outcomes were assessed using a histological scoring system. The 2.5 mm burr method was subsequently examined in 4 live rabbits (one eye per animal). In one animal the peripheral limbus was left intact. The clinical appearance of each eye was monitored for changes in clarity, vascularization and fluorescein staining for up to 5 weeks and followed by histology (H&E, PAS and immunostaining for cytokeratin 3 and cytokeratin 13).

Results : The 2.5 mm burr most reliably removed epithelial cells from the cornea and limbus. In contrast, small islands of limbal epithelium were occasionally retained following limbectomy or application of the 1 mm burr. Five weeks after in vivo application of the 2.5 mm burr to the entire cornea and limbus, all three eyes displayed neovascularization and retained epithelial defects of ~ 40%. While goblet cells were only observed by PAS staining in 2 eyes, the presence of conjunctival epithelial cells was confirmed in all 3 eyes by immunohistochemistry for cytokeratin 13. By comparison, the single eye with an intact limbus appeared to heal normally within 2 weeks (no corneal vascularization, no goblet cells and positive staining for cytokeratin 3).

Conclusions : The Algerbrush II rotating burr is an effective tool for induction of LSCD in New Zealand White rabbits. Moreover, best results are achieved using the 2.5 mm burr. We propose that the efficiency of this new method will reduce the number of animals required during development of new treatments for LSCD.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.