Administration of Nitric Oxide Through a Novel Copper-Chitosan Delivery System in Human Corneal and Limbal Epithelial...

Nitric oxide (NO) has gained attention for its role in facilitating wound healing by promoting cell migration, while being cytoprotective in a variety of cell types. We determined the efficacy of NO, administered using a novel application of copper-chitosan treatments (Cu-Ch), in facilitating corneal epithelial wound healing using an in vitro model of corneal epithelial and limbal epithelial cell injury.

Human corneal epithelial (HCE) and human limbal epithelial (HLE) cells were monitored under no-scratch (CON), untreated scratch (CS), scratch + plain chitosan composite (0%), scratch + 1% copper solution Cu-Ch (1%), and scratch + 2% copper solution Cu-Ch (2%) conditions. Cell migration, cytotoxicity, apoptosis, and total nitrate/nitrite concentrations were measured at 24, 48, and 72 hours after injury and treatment. iNOS expression in HLE cells also was determined using Western blot.

Wound closure significantly increased in HCE cells treated with Cu-Ch (1% and 2%) after 72 hours, while HLE cells showed a significant decrease in closure with Cu-Ch (1% and 2%) treatment compared to CS. Cytotoxic fragments decreased significantly with 1% and 2% Cu-Ch treatments in HCE cells. Nitrate/nitrite levels in HLE cells showed a significant increase with 2% Cu-Ch treatment compared to CS. This increase is complemented with an upregulation of iNOS.

Overall, HCE wound healing was accelerated with administration of Cu-Ch treatment. Differences between HCE and HLE responses may be due to intrinsic differences in NO metabolism, as evidenced by differences in NO production, potentially caused by differences in iNOS expression with treatment.