An investigation into corneal enzymatic resistance following epithelium-off and epithelium-on corneal cross-linking protocols

The aim of this study was to investigate corneal enzymatic resistance following epithelium off and on riboflavin/UVA cross-linking (CXL). One hundred and fourteen porcine eyes were divided into four non-irradiated control groups and seven CXL groups. The latter comprised; (i) epithelium-off, 0.1% iso-osmolar riboflavin, 9 mW UVA irradiation for 10 min, (ii) disrupted epithelium, 0.1% hypo-osmolar riboflavin, 9 mW UVA for 10 min, (iii) epithelium-on, 0.25% hypo-osmolar riboflavin with 0.01% benzylalkonium chloride (BACS), 9 mW UVA for 10 min, (iv) epithelium-on, 5 min iontophoresis at 0.1 mA for 5 min with 0.1% riboflavin solution, 9 mW UVA for 10 min or (v) 12.5 min, (vi) epithelium-on, prolonged iontophoresis protocol of 25 min with 1.0 mA for 5 min and 0.5 mA for 5 min with 0.25% riboflavin with 0.01% BACS, 9 mW UVA for 10 min or (vii) 12.5 min. Enzymatic resistance was assessed by daily measurement of a corneal button placed in pepsin solution and measurement of corneal button dry weight after 11 days of digestion. This study revealed that the enzymatic resistance was greater in CXL corneas than non-irradiated corneas (p < 0.0001). Epithelium-off CXL showed the greatest enzymatic resistance (p < 0.0001). The prolonged iontophoresis protocol was found to be superior to all other trans-epithelial protocols (p < 0.0001). A 25% increase in UVA radiance significantly increased corneal enzymatic resistance (p < 0.0001). In conclusion, although epithelium-on CXL appears to be inferior to epithelium-off CXL in terms of enzymatic resistance to pepsin digestion, the outcome of epithelium-on CXL may be significantly improved through the use of higher concentrations of riboflavin solution, a longer duration of iontophoresis and an increase in UVA radiance.