Interfacial properties of corneal epithelial cells at different degrees of differentiation

Abstract

 

Purpose

 

Ocular surface disorders are a heterogeneous group of diseases that adversely affect the biochemical and biophysical properties of the corneal surface and the maintenance of a cohesive tear film. The corneal and conjunctival epithelia possess intricate topographical structures (microvilli and microplicae) covered by a thick glycocalyx (composed mainly of membrane-spanning mucins), which influence its wettability. The basic interfacial properties (e.g. surface energy) of the ocular surface and their relation to the presence of topography and/or secreted mucins are understudied. The purpose of this study was to determine the relation of mucin expression (MUC1/4/16) with the interfacial properties of corneal epithelial cells.

 

Methods

 

Limbal-derived immortalized human corneal epithelial cells (hTCEpi) were routinely cultured in growth medium (Epilife® + Epilife defined growth supplement). Following 100% confluence, Epilife was replaced with Stratification media (DMEM/F12 supplemented with 10 ng/mL EGF and 10% FBS) for up to 7 days to induce differentiation, stratification and mucin expression. Quantitative PCR was performed to determine mRNA expression of mucins at different stages of stratification. Topographical characterization of cell surface during differentiation was performed using AFM. Surface energy of the cells at the various stages of stratification was quantified by measuring the contact angle between a series of fluorocarbons immersed in media.

 

Results

 

Mucin expression was significantly elevated in cells cultured in stratification media with maximal expression observed at day 2-3 before decreasing to 50% maximum at day 7. Cells cultured in Epilife showed cobblestone-like morphology with planar cell membranes, while stratified cells had a flattened morphology and were heterogeneous, with differing levels of microvillus formation. The Lewis acid component of the surface energy of differentiated cells showed a 1.9 fold increase with respect to undifferentiated cells.

 

Conclusions

 

Mucin expression was increased and surface features developed upon stratification and this event may play a role in determining the interfacial properties of corneal epithelial cells. We speculate that modulating the surface chemistry and topography of the ocular surface points to novel pathways for the development of therapeutic strategies for the treatment of ocular surface disorders.

 

Keywords: 486 cornea: tears/tear film/dry eye • 485 cornea: surface mucins • 482 cornea: epithelium