Bioengineered keratocyte spheroids fabricated on chitosan coatings enhance tissue repair in a rabbit corneal stromal defect model

Cultivated cell spheroid transplantation is widely studied as a means of facilitating tissue regeneration. Chitosan biomaterial has been shown to promote keratocyte aggregation and multicellular spheroid formation. This study provides further evidence on application of bioengineered keratocyte spheroids for corneal stromal tissue engineering. In an allogeneic rabbit model of stromal destruction caused by bacterial keratitis, the corneas were intrastromally injected with isolated keratocyte suspensions or aggregated spheroid grafts at same cell number. Results of clinical observations and histological examinations on postoperative day 14 showed that when an antibiotic eye drop is only medication for inhibiting bacterial growth, permanent damage to stroma occurs, leading to disorganization of collagen lamellae and tissue structure as well as loss of corneal transparency and visual function. Intrastromal grafting of keratocytes provided additional benefits to overcome drawbacks of limited disease treatment performance associated with topically applied antibiotics. In particular, as compared to their cell suspension counterparts, bioengineered keratocyte spheroids had higher ability to preserve cellular phenotype, secrete collagen matrix, and enhance graft retention, suggesting excellent repair capability for managing stromal tissue defect and alleviating corneal haze/oedema. In summary, the findings emphasize the role of keratocyte configuration (i.e., two-dimensional monolayer or three-dimensional spheroid) in determining therapeutic potency of cellular allografts for stromal tissue reconstruction. Transplantation of keratocyte spheroids cultured on chitosan substrates may represent a promising strategy for corneal stromal repair.