Potential Functional Restoration of Corneal Endothelial Cells in Fuchs Endothelial Corneal Dystrophy by ROCK Inhibitor (Ripasudil).

PURPOSE: Rho-associated kinase (ROCK) inhibitors have been successfully used as a rescue strategy in eyes that failed to clear after descemetorhexis without endothelial graft for treatment of Fuchs endothelial corneal dystrophy (FECD). The functional mechanisms by which ROCK inhibitors modulate corneal endothelial cell regeneration in FECD patients have, however, not been clarified. Here, we analyzed the effect of the ROCK inhibitor ripasudil on corneal endothelial cells of FECD patients and normal donors using ex vivo tissue and in vitro cellular models. DESIGN: Experimental study: laboratory investigation. METHODS: This institutional study used endothelial cell-Descemet membrane lamellae from FECD patients (n = 450) undergoing Descemet membrane endothelial keratoplasty (FECD ex vivo model), normal research-grade donor corneas (n = 30) after scraping off central endothelial cells (ex vivo wound healing model), normal donor corneas (n = 20) without endothelial injury, and immortalized cell lines (n = 3) generated from FECD patients (FECD in vitro model). Descemet membrane lamellae were dissected into halves and incubated for 24-72 hours in storage medium with or without a single dose of 30 µM ripasudil. The effects of ripasudil on expression of genes and proteins related to endothelial cell proliferation, migration, functionality, and endothelial-to-mesenchymal transition were analyzed and complemented by functional assays on FECD cell lines. RESULTS: A single dose of ripasudil induced significant upregulation of genes and proteins related to cell cycle progression, cell-matrix adhesion and migration, as well as endothelial barrier and pump function up to 72 hours, whereas classical markers of endothelial-to-mesenchymal transition were downregulated in both FECD and normal specimens compared to unstimulated controls ex vivo. In addition to stimulation of proliferation and migration, ripasudil-induced changes in expression of functional signature genes could be also verified in FECD cell lines in vitro. CONCLUSIONS: These data support the concept that inhibition of ROCK signaling represents a potent tool in regenerative therapies in FECD patients through reactivation of cell proliferation and migration as well as restoration of endothelial pump and barrier function without inducing adverse phenotypic changes.

Preservation of the Corneal Epithelium in Different Corneal Storage Media.

PURPOSE: With increasing time, epithelial defects (EDs) develop in virtually all corneas stored in corneal storage media. Optisol GS and Life 4°C are commonly available intermediate storage media used for corneal storage before keratoplasty. Epithelial preservation capabilities of Life 4°C and Optisol GS are compared in this study. METHODS: Nine pairs of human corneas were harvested, and 1 cornea of each pair was stored in Optisol GS and the other was stored in Life 4°C. The size and frequency of EDs of corneas stored in Optisol GS and Life 4°C were measured over time within the chambers using a backlit approach for 14 to 17 days of storage. RESULTS: At poststorage days 4, 8, and 12, there were no statistical differences in the percent change in the area of the ED between both groups. Of corneas without initial EDs, 6 of 7 (85.7%) stored in Optisol GS and 5 of 8 (62.5%) stored in Life 4°C developed an ED by the end of the assessment period. At the end of the observation period, there was no significant difference in the change in the percent area of the ED between corneas stored in Optisol GS and Life 4°C [4.3% ± 6.6% and 2.1% ± 2.6%, respectively (P = 0.38)]. CONCLUSIONS: Optisol GS and Life 4°C storage media did not significantly differ in their abilities to preserve the corneal epithelium of the donor tissue for up to 17 days. Most corneas stored in both cold-storage media developed EDs within the 14-day observation period.