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J. L. Ubels, M. J. Lim, R. K. Hurst, B. J. Konynenbelt; Cytotoxicity Testing of Multi-Purpose Contact Lens Solutions Using Monolayer and Stratified Cultures of Human Corneal Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2009;50(13):6356.
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© ARVO (1962-2015); The Authors (2016-present)
To evaluate validity of in vitro cytotoxicity methods by measuring the effects of multi-purpose contact lens care solutions (MPS) on the morphology, viability, proliferation and barrier function of monolayer and stratified cultures of human corneal-limbal epithelial (HCLE) cells.
MPS tested: OPTI-FREE (OF), OPTI-FREE Express (OFE), ReNu Multiplus (RN), Complete Multipurpose (CM). Cells were exposed to the MPS for 10, 20 or 60 min. Monolayer cultures: Cell morphology following treatment and during 24 hr of recovery in culture medium was observed. Effects of MPS on cell viability were determined using a live/dead cell assay with flow cytometry. Cell plating efficiency and proliferation following exposure of monolayer cultures to MPS were evaluated. Stratified cultures: Cells were grown on permeable membrane supports. Cell morphology following treatment with MPS was observed. Effects of MPS on barrier function were determined by measuring fluorescein permeability and trans-epithelial resistance (TEER).
Monolayer cultures: All MPS caused time-dependent changes in morphology ranging from mild to moderate swelling and junction disruption at 10 min (OFE, CM) to marked swelling and complete breakdown of cell junctions at 60 min (OFE, RN, CM). Cells exposed to the MPS for 10 or 20 min recovered normal morphology within 24 hr; however, cells exposed to OFE for 60 min did not recover, and cells exposed to CM for 60 min remained swollen at 24 hr. OFE caused a 10-30% decrease in cell viability after 10-60 min, but OF, RN and CM had no effect. Exposure to MPS for 10 min did not affect cell proliferation. Exposure to RN or OFE for 20 or 60 min caused decreased plating efficiency when the cells were passed. All MPS caused significant decreases in cell proliferation rate after 20 or 60 min treatments. Stratified cultures: Morphology of stratified cells cultured on membrane supports and exposed to MPS did not change at 10, 20 or 60 min. None of the MPS caused an increase in fluorescein permeability or a decrease in TEER of stratified cultures.
Monolayer cultures are highly sensitive, with all MPS causing cytotoxicity. In contrast, none of the MPS tested damaged stratified HCLE cultures. It is suggested that models that simulate the stratified structure of the corneal epithelium should be used for in vitro toxicologic testing of contact lens care solutions. This will prevent collection of misleading data on clinically safe, beneficial products.
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