Confocal microscopy and histopathological examination of diffuse lamellar keratitis in an experimental animal model

Abstract

Purpose: To investigate the cell populations and structural alterations of the cornea in an experimental model of diffuse lamellar keratitis (DLK) using confocal microscopy and histopathology.

Methods: A corneal flap was cut in 22 eyes of 11 New Zealand rabbits and the stromal interface was exposed to balanced salt solution (BSS, BSS group) and Pseudomonas aeruginosa lipopolysaccharide (LPS) endotoxin (5 mg/mL) (LPS 5 mg/mL group) and (3.5 mg/mL) (LPS 3.5 mg/mL group). Postoperatively, eyes were examined with a slit-lamp microscope (DLK grading) and confocal microscopy. Animals were sacrificed on day 3 (BSS group and LPS 5 mg/mL group) and day 4 (LPS 3.5 mg/mL group). Corneoscleral buttons were excised and processed for histopathologic examination.

Results: Seven eyes were excluded. Slit-lamp microscopy revealed no cellular infiltration in the BSS group (five eyes). In the LPS groups, all eyes developed DLK, with iritis only observed in grade III eyes. In the LPS 5 mg/mL group, four eyes had DLK grade III, with iritis in three eyes. In the LPS 3.5 mg/mL group, three eyes had grade II and three eyes had grade III with iritis. On confocal microscopy, the BSS group had no cellular infiltration. Dense accumulation of inflammatory cells at the interface was noted in both LPS groups. Histopathology in the BSS group had a normal appearance. In the LPS groups, an inflammatory infiltrate was present at the interface that consisted of three cell populations--eosinophils, neutrophils, and lymphocytes.

Conclusions: Lipopolysaccharide endotoxin induced DLK in all exposed eyes, with iritis in a considerable proportion of eyes. The infiltrate consisted of three cell populations. Confocal microscopy showed the infiltrate in all affected eyes. Histopathological and confocal microscopic findings correlated well with the clinical appearance.