These results indicate that using cultured HCECs and functional biomaterials, PNIPAAm and gelatin, an effective cell sheet-based therapy can be developed for the treatment of corneal endothelium deficiency.

IntroductionDry-eye syndrome (DES) is a general eye disease. Eye drops are the common ophthalmological medication. However, the ocular barrier makes it difficult to attain high drug bioavailability. Nanomedicine is a promising alternative treatment for ocular diseases and may increase drug content in the affected eye.MethodsTo explore this potential, we constructed nanoparticles (NPs) containing an anti-inflammatory agent for DES treatment. The NPs were made of gelatin–epigallocatechin gallate (EGCG) with surface decoration by hyaluronic acid (HA) and designated “GEH”. The particle size, surface charge, and morphology were evaluated. The in vitro biocompatibility and anti-inflammation effect of nanoparticles were assayed via culturing with human corneal epithelium cells (HCECs) and in vivo therapeutic effect was examined in a DES rabbit’s model.ResultsThe synthesized GEH NPs had a diameter of approximately 250 nm and were positively charged. A coculture experiment revealed that 20 µg/mL GEH was not cytotoxic to HCECs and that an EGCG concentration of 0.2 µg/mL downregulated the gene expression of IL1B and IL6 in inflamed HCECs. Large amounts of GEH NPs accumulated in the cytoplasm of HCECs and the ocular surfaces of rats and rabbits, indicating the advantage of GEH NPs for ocular delivery of medication. Twice-daily topical treatment with GEH NPs was performed in a rabbit model of DES. The ocular surface of GEH-treated rabbits displayed normal corneal architecture with no notable changes in inflammatory cytokine levels in the cornea lysate. The treatment improved associated clinical signs, such as tear secretion, and fluorescein staining recovered.ConclusionWe successfully produced GEH NPs with high affinity for HCECs and animal eyes. The treatment can be delivered as eye drops, which retain the drug on the ocular surface for a longer time. Ocular inflammation was effectively inhibited in DES rabbits. Therefore, GEH NPs are potentially valuable as a new therapeutic agent delivered in eye drops for treating DES.

Cell transplantation strategies usually involve the use of supporting carrier materials because of the soft and fragile nature of these grafts. In this work, a cell-adhesive gelatin hydrogel carrier was fabricated to deliver cultivated human corneal endothelial cell (HCEC) sheets, which were harvested from thermo-responsive poly(N-isopropylacrylamide) (PNIPAAm)-grafted culture surfaces. The carrier disks, consisting of gelatins with a different isoelectric point (IEP = 5.0 and 9.0) and a molecular weight (MW) ranging from 3 to 100 kDa, were subjected to 16.6 kGy gamma irradiation for sterilization. The effect of IEP and MW of the raw gelatins (i.e., before irradiation) on the functionality of sterilized disks was studied by determinations of mechanical property, water content, dissolution degree, and cytocompatibility. Irrespective of the IEP of raw gelatin, hydrogel disks prepared with high MW (100 kDa) exhibited a greater tensile strength, lower water content, and slower dissolution rate than those made of low MW gelatin (8 and 3 kDa). From the investigation of cellular responses to the disks, the negatively charged gelatin (IEP = 5.0) groups were more cytocompatible when compared with their positively charged counterparts (IEP = 9.0) at the same MW (100 kDa). Additionally, in the negatively charged gelatin groups, only a slight increase in pro-inflammatory cytokine expression was observed with increasing MW of gelatin from 3 to 100 kDa. It is concluded that the gamma-sterilized hydrogel disks made from raw gelatins (IEP = 5.0, MW = 100 kDa) with appropriate dissolution degree and acceptable cytocompatibility are capable of providing stable mechanical support, making these carriers promising candidates for intraocular delivery of cultivated HCEC sheets.