Abstract
Purpose :
Dry eye disease (DED) is characterized by disruption in tear film homeostasis involving increased inflammation, epithelial barrier dysfunction, and ocular surface damage. Adherens junctions, which are essential for maintaining epithelial barrier integrity, are formed by cadherins and catenins, which also contribute to the Wnt/β-catenin signaling pathway that regulates cell proliferation and migration during inflammation. Using an in vitro model of DED, we hypothesize that increased hyperosmolarity alters expression of cadherin and catenins, leading to impaired barrier function and wound healing via changes to Wnt/β-catenin activation.
Methods :
Telemorase-immortalized human corneal epithelial (hTCEpi) cells cultured in normal osmolar medium (312 mOsM) were treated with 70, 90, and 120 mM NaCl for 24 hours to achieve 450, 500, and 550 mOsM, respectively. Epithelial barrier function and cell migration (wound healing) were measured by electric cell-substrate impedance sensing (ECIS) apparatus, and E-cadherin, β-catenin, and p120 catenin levels were assessed using RT-PCR, western blotting, and immunofluorescence staining.
Results :
Hyperosmolarity significantly decreased trans-epithelial resistance (TER) in a dose-dependent manner in hTCEpi cells, with ~30% decline observed in the 500 and 550 mOsM groups after 48 hours. Similarly, wound healing assay showed significant decrease in cell migration rate in all treatment groups 48 hours post-wounding (untreated, 450, 500, and 550 mOsM recorded 21.7 µm/hr, 7.9 µm/hr, 7.3 µm/hr, and 0 µm/hr migration rates, respectively). Furthermore, hyperosmolarity significantly decreased expression of E-cadherin (0.44-fold, p=0.0026), β-catenin (0.38-fold, p=0.0027), and p120-catenin (0.37-fold, p=0.0202).
Conclusions :
In conclusion, our results indicate that epithelial barrier function and wound healing were significantly altered by hyperosmolarity in a dose-dependent manner, and these changes were accompanied by a reduction in the expression of adherens junction proteins, E-cadherin and β-catenin. Taken together, these data provide evidence impairing Wnt/β-catenin signaling in DED. Future experiments targeting this pathway will prove useful in identifying potential therapeutic targets for DED.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.