Riboflavin Concentration Analysis in Rabbit Corneas Before and After Corneal Collagen Cross-Linking Using Confocal Laser Scanning Microscopy.

Abstract

The aim of this study was to evaluate riboflavin in rabbit corneas before and after corneal collagen cross-linking (CXL) using confocal laser scanning microscopy. A randomized controlled experimental study. The study was divided into 3 parts. In part 1 of this study, 30 eyes from 15 rabbits were used to evaluate riboflavin in de-epithelialized corneas before standard CXL. In part 2, 12 eyes from 6 rabbits were used to compare the differences of riboflavin concentration between standard CXL and transepithelial CXL (TE-CXL). In part 3, 12 eyes from another set of 6 rabbits were used to evaluate 1-day postoperative changes of riboflavin concentration between standard CXL and TE-CXL. Riboflavin concentrations in corneas were evaluated by calculating fluorescence densities with a confocal laser scanning microscope. In part 1, the riboflavin concentration after a 20-minute instillation was 0.036%, and did not reach the safety threshold (0.040%) for standard CXL. In part 2, the riboflavin concentration before TE-CXL was lower than 0.010%, which was significantly lower than standard CXL (P < 0.001). In part 3, corneal fluorescence decreased by approximately 100% (fluorescence quenching) 1 day after standard CXL and approximately 60% 1 day after TE-CXL. Application of 0.010% riboflavin solution for 30 minutes is essential for standard CXL. TE-CXL is not recommended because the epithelial layers are inadequately permeable to riboflavin during and 1 day after surgery.