Cells
The Effects of Rho-Associated Kinase (ROCK) Inhibitor Supplementation in Corneal Preservation Media for Donor Tissue Used in Descemet’s Membrane Endothelial Keratoplasty (DMEK)
By Hon Shing Ong, Deepika Malhotra, Dawn Jing Hui Neo and 4 others.
Abstract
Purpose
This study seeks to evaluate the effects of pre-treating donor corneal tissue with a Rho-associated kinase inhibitor (ROCKi), Netarsudil (AR-13324), on the viability of corneal endothelial cells (CECs) during Descemet’s membrane endothelial keratoplasty (DMEK) graft preparation and insertion using endothelium-in and endothelium-out techniques.
Methods
This ex vivo comparative study utilized 24 pairs of research-grade human donor corneas preserved in Optisol-GS. One cornea from each pair was incubated in 1 µM AR-13324 for 2 h, while the contralateral cornea served as a control. DMEK tissues were prepared and simulated insertion was performed using either the endothelium-in (Endoglide) or endothelium-out (Geuder glass injector) technique. The surgeon performing the tissue preparation and insertion was masked to the ROCKi treatment allocation. Post-manipulation, tissues underwent live/dead cytotoxicity assay using Calcein-AM and Ethidium homodimer-1 staining. Percentages of live, fallout, and dead CECs were quantified via fluorescence microscopy and ImageJ (version 1.54p) analysis. Correlations between donor age, preservation duration, and cell viability were also analyzed.