| Corneal transplantation is considered the definitive treatment for corneal blindness, which affects more than 10 million people worldwide. However, declining donor tissue availability and the risk of donor graft rejection motivate the search for viable alternatives. The overall goal of this research was to develop strategies for ocular surface reconstruction and in vitro corneal modeling. To that end, collagen vitrigel (CV), a micron-scale, transparent, collagen film was evaluated for corneal repair and a novel bioadhesive, chondroitin sulfate (CS) crosslinked with poly(ethylene glycol)-amine (PEG-(NH 2)6) (CS-PEG) was examined for sealing corneal wounds. An in vitro corneal microtissue model comprised of a CV film reversibly incorporated into a microfluidic device has also been developed.In vitro evaluations of limbal stem cell (LSC) cultures on CV resulted in partially stratified epithelial cell sheets with upregulation of stem cell maker, p63, compared to collagen coated plates. LSCs were first expanded on CV, followed by the transplantation of confluent cell sheets into a rabbit model of limbal insufficiency. Compared to negative control, corneal clarity was much improved, while neovascularization was less in the transplanted group. In a second in vivo study, the goal was to assess the efficacy of anterior corneal reconstruction using fibrin glue (FG)-assisted acellular CV transplantation in an animal model of deep stromal injury. All corneas receiving precut CV + FG remained optically clear through 10 weeks and demonstrated stable host-graft integration.Histological analysis of the CS-PEG adhesive in mouse, swine and rabbit models demonstrated that the adhesive is effective in bridging a corneal wound, is biodegradable, induces minimal inflammatory response, resists epithelial cell ingrowth and does not induce scarring.Microfludic devices were fabricated in which CV was used both as a functional and sacrifical cell growth substrate for the culture of corneal microtissue patches. The utility of the platform was demonstrated by measuring on-chip transepithelial permeability.These studies highlight the versatility of CV as a tool for ocular surface reconstruction and in vitro modeling. These results suggest that CS-PEG may not only be used as a corneal wound sealant, but also in combination with CV for several applications in corneal repair. |
