Glycerol Modulated Collagen Fibril Evolution and Lamellar Organization for Biomimetic Corneal Substitutes Construction

Collagen as the main structural component of the cornea exhibits unique and highly organized fibril lamellae, which contribute to the maintenance of corneal structure and transparency. Nevertheless, collagen assembly in vitro to create ideal artificial corneal substitutes with human cornea comparable thickness and optics is still limited. Here, glycerol as a regulator can reconcile collagen thickness, transparency, and permeability, a conflicting goal by current keratoprosthesis strategies. Structure analysis reveals that glycerol treatment induces collagen hydrogels to undergo a sequential three-step multiscale structural evolution: weakened collagen crystallization at the molecular level, followed by ordered and distanced microfibril packaging at the nanoscale, and ultimately lamellar structure as well as fibril diameter and spacing-dependent optics at a macroscopic level. Such ultrastructure is then stabilized by oxazolidine crosslinking to obtain a collagen-based artificial corneal substitute (Col-Gly-OX) with optimal integration of optical clarity, mechanical robustness, high permeability, manufacturability, easy preservation and in vitro biocompatibility. Further in vivo study demonstrates that Col-Gly-OX displays excellent tissue integration, epithelialization, and stromal remodeling in a rabbit lamellar keratectomy. Overall, this work illustrates the potential of glycerol regulator to mediate the multiscale structural organization of collagen, providing a green, simple and effective strategy for the development of bionic artificial cornea.