Abstract
Purpose :
The use of topical Rho-kinase (ROCK) inhibitors for treatment of corneal endothelial disease is expanding rapidly, but uncertainty remains regarding their differential effects on corneal endothelial and epithelial cells. Here, we analyzed the effects of ROCK inhibitors ripasudil and netarsudil on expression of function-related markers in corneal endothelial and epithelial cells in vitro.
Methods :
Primary human corneal endothelial and epithelial cells were incubated with ripasudil (10 and 30 µM) and netarsudil (1 µM) for up to 7 days. The effects of ROCK inhibitors on expression of genes and proteins related to endothelial and epithelial cell function were analyzed by qRT-PCR and immunocytochemistry and complemented by functional assays (Na+/K+-ATPase activity assay, TEER assay, FITC-Dextran permeability assay, BrdU proliferation assay, migration assay).
Results :
In corneal endothelial cells, both ripasudil and netarsudil significantly stimulated cell proliferation and migration, and increased Na+/K+-ATPase activity and barrier function compared to untreated controls. Both drugs further induced significant upregulation of genes and proteins related to pump and barrier function including Na,K-ATPase subunits, bicarbonate transporter, monocarboxylate transporter, chloride transporter, aquaporins, and tight junction proteins. In contrast, differential effects of both ROCK inhibitors were observed on expression of corneal epithelial cell junctions: Whereas expression levels of adherens junctions, (hemi)desmosomes and tight junctions were consistently upregulated by ripasudil, they were variably downregulated by netarsudil along with a corresponding decline in epithelial barrier function. Expression changes normalized upon discontinuation of netarsudil.
Conclusions :
The findings suggest that both ripasudil and netarsudil can improve corneal endothelial cell function, but exert differential effects on corneal epithelial cell junctions. Reversible bullous epithelial edema, occasionally reported after the use of netarsudil, may be caused by direct effects on corneal epithelial cell-cell and cell-basement membrane junctions.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.