Corneal Enzymatic Resistance Following High-Fluence Accelerated Corneal Cross-Linking With Riboflavin And Uv-A Ex Vivo

Classic corneal cross-linking (CXL) uses UV-A light and riboflavin to generate reactive oxygen species (ROS) in the stroma. ROS not only kill pathogens, but also bind together stromal molecules – strengthening the cornea and hiding collagenase binding sites, increasing resistance to enzymatic digestion. In infectious keratitis, reduced pathogen-induced digestion equals less melting and smaller ultimate scar sizes. We compared the digestion-inhibiting effects of traditional CXL protocols with newer higher fluence protocols.

Laboratory investigation. ELZA Institute, Dietikon, Switzerland and Dept. of Ophthalmology, University of Antwerp, Antwerp, Belgium

Ex vivo porcine corneas were assigned to 5 groups. Groups 1 and 2 were controls (abrasion only and abrasion/riboflavin soaking, respectively). Group 3 received CXL treatment using a slightly accelerated low-fluence cross-linking protocol (5.4 J/cm², 10′ @ 9 mW/cm²), whereas groups 4 and 5 were treated with accelerated high-fluence protocols (10 J/cm², 9′15″ @ 18 mW/cm² and 15 J/cm², 8′20″ @ 30 mW/cm², respectively). CXL procedures were performed using 0.1% riboflavin (Ribo-Ker, EMAGine SA, Zug, Switzerland) and a commercially available CXL device (C-eye, EMAGine AG, Zug, Switzerland). All corneas were digested in 0.3% collagenase A solution. Mean time until complete dissolution was determined.

Both low-fluence and high-fluence cross-linking protocols significantly increased resistance to digestion. The mean times to digestion in Groups 1 through 5 were: 21.4 ± 3.25 h, 22.3 ± 1.97 h, 26.5 ± 3.54 h, 30.6 ± 1.79 h, and 32.3 ± 2.56 h, respectively. Groups 3–5 displayed significantly higher (p<0.001) digestion resistance than either control group (Groups 1 and 2). The accelerated, high-fluence protocols of Groups 4 and 5 significantly (p<0.001) rendered corneas more digestion-resistant than those in the 5.4 J/cm²-fluence Group 3. The times to complete dissolution were not significantly different between groups 4 and 5 (p<0.65).

High-fluence photoactivated chromophore for keratitis-CXL (PACK-CXL) protocols (10 and 15 J/cm²) are more effective at treating infectious keratitis than lower fluence, traditional corneal ectasia treatment protocols (5.4 J/cm²). The study shows that high-fluence CXL protocols can significantly increase resistance to enzymatic digestion, but that this effectively plateaus at fluences of 10 J/cm². These data support and reinforce that higher fluence PACK-CXL, in addition to the already established and published higher pathogen killing effect, also should result in less corneal melting and smaller scar sizes than traditional 5.4 J/cm² protocols.