Chondroitin Sulfate-Based Biocompatible Crosslinker Restores Corneal Mechanics and Collagen Alignment

Invest Ophthalmol Vis Sci. 2017 Aug 1;58(10):3887-3895. doi: 10.1167/iovs.16-21292.


Purpose: To evaluate the crosslinking effect of functionalized chondroitin sulfate (CS) in an ex vivo rabbit cornea model.

Methods: Chondroitin sulfate molecules were chemically modified with the N-hydroxysuccinimide (NHS) group. Enucleated rabbit eyes were crosslinked with 2, 5, or 10 mg/mL CS-NHS solution for 30 or 60 minutes. The CS-NHS penetration, corneal swelling ratio, Young's modulus, and ultrastructure of the crosslinked corneas were characterized. In addition, rabbit corneas were further treated with a collagenase-chondroitinase solution to create an ex vivo keratoconus (KC)-like model. The KC model corneas were crosslinked with a standard riboflavin-ultraviolet (UV) method or alternatively with CS-NHS. Corneal mechanics, ultrastructure, and keratocyte gene expression were evaluated after UV and CS-NHS crosslinking.

Results: CS-NHS effectively penetrated into the corneal stroma within 60 minutes of treatment initiation. CS-NHS crosslinking reduced the swelling ratio by 35%, increased Young's modulus by 20%, and increased collagen fibril diameter and density. CS-NHS crosslinking improved corneal mechanics of KC model corneas to levels comparable to those with UV crosslinking. Moreover, CS-NHS crosslinking demonstrated significant downregulation of proinflammatory gene expression of keratocytes, indicating a potential protective effect imparted by CS-NHS during crosslinking.

Conclusions: Our results demonstrated that CS-NHS can reinforce normal and KC model corneal mechanics, and restore collagen density and alignment in KC model corneas without causing extensive keratocyte apoptosis and proinflammatory gene upregulation. Therefore, CS-NHS crosslinking can potentially provide an effective, safe, and biocompatible means of corneal reinforcement.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Biomechanical Phenomena
  • Chondroitin Sulfates / pharmacology*
  • Collagen / metabolism*
  • Cornea / drug effects*
  • Cornea / metabolism
  • Cornea / physiopathology
  • Corneal Keratocytes / drug effects
  • Corneal Keratocytes / metabolism
  • Cross-Linking Reagents / pharmacology*
  • Disease Models, Animal
  • Elastic Modulus / drug effects
  • Photosensitizing Agents / pharmacology
  • Rabbits
  • Ultraviolet Rays


  • Cross-Linking Reagents
  • Photosensitizing Agents
  • Chondroitin Sulfates
  • Collagen