Transepithelial Riboflavin Absorption in an Ex Vivo Rabbit Corneal Model

• Published in: Investigative ophthalmology & visual science 2015-07, Vol.56 (8), p.5006-5011
• Main Authors: Gore, Daniel M, O'Brart, David, French, Paul, Dunsby, Chris, Allan, Bruce D
• Format: Article
• Language: English
• Subjects: Animals; Corneal Stroma - metabolism; Corneal Stroma - pathology; Disease Models, Animal; Epithelium, Corneal - metabolism; Epithelium, Corneal - pathology; Keratoconus - drug therapy; Keratoconus - metabolism; Keratoconus - pathology; Microscopy, Fluorescence; Ophthalmic Solutions; Rabbits; Riboflavin - administration & dosage; Riboflavin - pharmacokinetics; Vitamin B Complex - administration & dosage; Vitamin B Complex - pharmacokinetics
• ISSN: 1552-5783; 1552-5783
• DOI: 10.1167/iovs.15-16903

To measure depth-specific riboflavin concentrations in corneal stroma using two-photon fluorescence microscopy and compare commercially available transepithelial corneal collagen cross-linking (CXL) protocols. Transepithelial CXL riboflavin preparations--MedioCross TE, Ribocross TE, Paracel plus VibeX Xtra, and iontophoresis with Ricrolin+--were applied to the corneal surface of fresh postmortem rabbit eyes in accordance with manufacturers' recommendations for clinical use. Riboflavin 0.1% (VibeX Rapid) was applied after corneal epithelial debridement as a positive control. After riboflavin application, eyes were snap frozen in liquid nitrogen. Corneal cross sections 35-μm thick were cut on a cryostat, mounted on a slide, and imaged by two-photon fluorescence microscopy. Mean (SD) concentrations were calculated from five globes tested for each protocol. Peak riboflavin concentration of 0.09% (± 0.01) was observed within the most superficial stroma (stromal depth 0-10 μm) in positive controls (epithelium-off). At the same depth, peak stromal riboflavin concentrations for MedioCross TE, Ricrolin+, Paracel/Xtra, and Ribocross TE were 0.054% (± 0.01), 0.031% (0.003), 0.021% (± 0.001), and 0.015% (± 0.004), respectively. At a depth of 300 μm (within the demarcation zone commonly seen after corneal cross-linking), the stromal concentration in epithelium-off positive controls was 0.075% (± 0.006), while at the same depth MedioCross TE and Ricrolin+ achieved 0.018% (± 0.006) and 0.016% (0.002), respectively. None of the remaining transepithelial protocols achieved concentrations above 0.005% at this same 300-μm depth. Overall, MedioCross TE was the best-performing transepithelial formulation. Corneal epithelium is a significant barrier to riboflavin absorption into the stroma. Existing commercial transepithelial CXL protocols achieve relatively low riboflavin concentrations in the anterior corneal stroma when compared to gold standard epithelium-off absorption. Reduced stromal riboflavin concentration may compromise the efficacy of riboflavin/ultraviolet corneal CXL.