RESUMO
OBJECTIVE: To evaluate the effects of residual hydrogen peroxide (H2O2) in neutralized H2O2-based contact lens-disinfecting solutions on morphology, viability, and barrier function of monolayer and stratified human corneal-limbal epithelial (HCLE) cells. METHODS: Cells were exposed to contact lens formulations containing 0.01% H2O2 for 10, 20, or 60 minutes. The morphology of monolayer or stratified cells was observed by microscopy. Monolayer or stratified cell viability was determined using a live/dead assay, and monolayer cell viability was quantified using flow cytometry. Effects of formulations on barrier function of stratified HCLE cells were evaluated by measuring fluorescein permeability and transepithelial resistance of cultures grown on membrane inserts. To determine the sensitivity of the tests to peroxide damage, stratified cells were also exposed to 0.01% to 0.3% H2O2 in culture medium. RESULTS: All formulations caused swelling of monolayer cells. Formulations with or without H2O2 at pH 7.9 caused mild decreases in monolayer cell viability but did not affect viability or barrier function of stratified HCLE cells. H2O2 (≥0.1%) in culture medium caused damage without recovery to stratified HCLE cells. CONCLUSIONS: Although tests on stratified cells are capable of detecting damage caused by H2O2 in culture medium, residual H2O2 in neutralized ophthalmic formulations had no effect on stratified cells in vitro. These data suggest that H2O2, used appropriately, is a safe disinfectant. Data comparing monolayer and stratified cultures suggest that monolayers are more sensitive to peroxide damage and that the effects of neutralized formulations on stratified cells may better predict the intact corneal epithelial response.
