The effect of limbal tissue with conjunctival flap and bovine amniotic membrane on alkaline burn corneal wound: experimental study in rabbits

Corneal ulcers pose a significant threat to vision and require timely, effective intervention to prevent permanent damage. This experimental study evaluated the therapeutic potential of combining crushed limbal tissue with either bovine amniotic membrane (AM) or a conjunctival flap to enhance corneal wound healing in a rabbit model. Twenty-five New Zealand white rabbits were randomly assigned to five groups: (G1) untreated control, (G2) AM alone, (G3) conjunctival flap alone, (G4) AM with crushed limbal tissue, and (G5) conjunctival flap with crushed limbal tissue. Standardized central corneal alkaline burns were induced using 1N sodium hydroxide. Healing was monitored over six weeks through fluorescein staining, serial imaging, and histopathological analysis. By week six, epithelial defect areas were significantly reduced in G2 (p = 0.031) and G4 (p = 0.011) compared to controls. G3 showed moderate improvement, while G5 exhibited variable outcomes with delayed healing in some animals. Only G1 showed progressive epithelial deterioration over time (p = 0.039). All treated groups (G2–G5) demonstrated early neovascularization, with partial regression by week six. Histologically, keratocyte loss was significantly reduced in G4 and G5 (p < 0.004), and stromal inflammation was milder in G2 (p = 0.03). Retrocorneal membrane formation was most frequent in G3. Among all groups, G4 showed the most consistent regenerative outcomes. These findings support the application of bovine AM combined with autologous limbal tissue as a practical and biologically active strategy for corneal repair in veterinary models. Further studies are warranted to evaluate long-term efficacy and optimize integration methods.