Measurements of Mucin Growth and Removal in Human Corneal Epithelia

Thomas Angelini

Abstract

Purpose:

Contact lens (CL) comfort has been correlated to lens friction coefficient. However, our understanding of the underlying mechanisms has been impeded by the difficulty of making real-time observations of eye-lens interactions at the cellular level. The purpose of this study is to establish an in vitro method to study in detail the interaction between cells and CL (or lens-like materials). Such an approach couples live-cell imaging to microtribometry and requires protocols for quantifying the rate at which corneal epithelial cells produce a protective mucin layer, enabling measurements of the living mucin-coated monolayer’s sensitivity to friction forces.

Methods:

Immortalized human corneal epithelial cells (hTCEpi) were cultured on fibronectin-coated glass-bottomed petri dishes. Mucin was imaged using a fluorescent carbohydrate stain and cell bodies dyed with calcein-Red. Time-lapse imaging was performed on an inverted confocal microscope equipped with a stage-top incubator. To shear the mucin and the cells under direct contact sliding, a microtribometer system was mounted onto an inverted microscope. With this system, normal and shear forces were applied to hTCEpi monolayers by a polyethylene-glycol hydrogel hemisphere while collecting fluorescence images of the living cells.

Results:

The rate at which the cells produce the mucin monolayer was measured. Quantitative image processing of the confocal z-stacks collected in time-lapse revealed the thickness of the mucin layer increasing at a rate of 2 µm/day. Our tribological sliding tests showed that mucin is not physically removed at low contact pressures. At high contact pressures >20 kPa, the mucin layer is erased and cell damage observed.

Conclusions:

CL lubricity in the eye is driven by lens interactions with tissue and tear film. The lens-tissue interface is very different from a lens-glass interface, which was shown using a friction probe that possesses material properties indicative of tissue.

Details

Year: 2016

Program Number: 160119

Author Affiliation: n/a

Co-Authors: Tristan Hormel, Juan Urueña, Angela Pitenis, Kyle Schulze, W. Sawyer

Co-Author Affiliation: University of Florida, University of Florida, University of Florida, University of Florida, University of Florida

Room: 211AB