Abstract
Purpose :
Ion channels are critical regulators of fluid hydration in many parts of the body. Channels such as CFTR and ENaC help maintain the quantity of fluid on the eye. As such, these channels are intriguing targets for the treatment of dry eye disease. This work investigates the ability of novel ENaC inhibitory peptides to regulate fluid dynamics in cultured rabbit and human corneal epithelial cells.
Methods :
Rabbit and human corneal epithelial cells were grown on differentiated on transwell membranes. Surface ENaC levels were determined by surface biotinylation and western blot analysis. Apical fluid height was assessed by confocal microscopy. Cell viability was determined by PrestoBlue staining. Cells were either untreated, treated with the classic ENaC inhibitor amiloride, or treated with one or more proprietary ENaC regulatory peptides.
Results :
Peptides were found to internalize ENaC subunits from the surface of rabbit and human corneal epithelial cells. This effect was not observed by amiloride, a classic small molecule inhibitor of ENaC. Internalization of ENaC was rapid, occurring in under 30 minutes, and sustained for at least eight hours after a single administration. Apical fluid height in peptide treated cells was significantly increased compared to amiloride or vehicle treatments. Finally, all peptides tested were found to be non-toxic without observed reductions in cell viability.
Conclusions :
ENaC inhibitory peptides safely internalized ENaC in human and rabbit corneal epithelial cells resulting in increased apical hydration. This data supports the continued development of ENaC inhibitory peptides for the treatment of dry eye disease.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.