3D printed biomimetic epithelium/stroma bilayer hydrogel implant for corneal regeneration

Corneal regeneration has always been a challenge due to its sophisticated structure and undesirable keratocyte-fibroblast transformation. Herein, we propose 3D printing of a biomimetic epithelium/stroma bilayer implant for corneal regeneration. Gelatin methacrylate (GelMA) and long-chain poly(ethylene glycol) diacrylate (PEGDA) are blended to form a two-component ink, which can be printed to different mechanically robust programmed PEGDA-GelMA objects by Digital Light Processing (DLP) printing technology, due to the toughening effect of crystalline crosslinks from long-chain PEGDA on GelMA hydrogel after photo-initiated copolymerization. The printed PEGDA-GelMA hydrogels support cell adhesion, proliferation, migration, meanwhile demonstrating a high light transmittance, and an appropriate swelling degree, nutrient permeation and degradation rate. A bi-layer dome-shaped corneal scaffold consisting of rabbit corneal epithelial cells (rCECs)-laden epithelia layer and rabbit adipose-derived mesenchymal stem cells (rASCs)-laden orthogonally aligned fibrous stroma layer can be printed out with a high fidelity and robustly surgical handling ability. This bi-layer cells-laden corneal scaffold is applied in a rabbit keratoplasty model. The post-operative outcome reveals efficient sealing of corneal defects, re-epithelialization and stromal regeneration. The concerted effects of microstructure of 3D printed corneal scaffold and precisely located cells in epithelia and stroma layer provide an optimal topographical and biological microenvironment for corneal regeneration.