Understanding the fate of allo-CLET, allo-SLET and CLAL in Alger brush-induced rabbit model of alkali burn

Abstract

Purpose : The current study aims to optimize mechanical LSCD generation and compare the fate of various allograft transplantation methods in epithelial regeneration.

Methods : Mechanical debridement of limbus was performed using the Alger brush-II (1.0-mm round-ended burr) after 360° limbal peritomy in New Zealand white rabbit (n=18) to generate LSCD. The eyes were observed for corneal opacity, vascularization and epithelial defect using clinical evaluation, slit lamp imaging, and anterior segment optical coherence tomography (AS-OCT). 4–8 weeks post debridement, Allogenic Simple Limbal Epithelial Transplantation (Allo-SLET), Conjunctival Limbal Allograft (CLAL) and Allogenic Cultivated Limbal Epithelial Transplantation (Allo-CLET) were performed with graft obtained from the Dutch-Belted rabbit. The transplanted eye was graded for vascularization, corneal opacity, and corneal epithelialization based on the scoring (1-5). Eyes were also assessed for central corneal epithelial thickness (CCET) changes The expression levels of inflammatory, corneal epithelial, keratinization, stem cell-specific and conjunctivalization markers were assessed using qRT-PCR and ELISA

Results : The severe and mild LSCD corneas exhibited opacity and vascularization scores, 2.33 ±0.33, 2.50 ± and 0.66±0.33, 1.33±0.03 respectively. Stable ocular surface and epithelial regeneration (CK3) were observed in all three transplantation methods. The LSCD cornea showed a significant decrease in corneal epithelial (CK12) and stem cell marker (ABCG2 & Np63α) expression (p≤0.0001, n=3) compared to the control eye. Allo-SLET and Allo-CLET treated eyes showed a significant increase (P= 0.0079 & 0.0019) in CCET compared to CLAL at 12 weeks of transplantation. The levels of tear cytokines (TNF-α, TGF-β, IL-4, IL-6, IL-10, and IL-17) didn’t exhibit any significant difference between Allo-SLET, Allo-CLET and CLAL. Our results show a stable ocular surface was achieved in all cases as there was no recurrent epithelial erosion post-transplantation

Conclusions : Alger brush-induced mechanical debridement is a suitable method for LSCD generation with persistent corneal epithelial defects. Different limbal allograft transplantation methods lead to stable ocular surface and comparable corneal epithelial restoration and thus confirm that there are no remarkable immunological differences in various allo-limbal transplantation.

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