Effect of S-Nitrosylation on post-mortem donor cornea endothelial health during hypothermic storage period

Abstract

Purpose : Protein S-nitrosylation (SNO) is the means by which nitric oxide (NO) bioactivity is conveyed throughout the body to regulate all manners of cellular function. Our previous work has suggested that addition of one or more SNO agents to preservation solutions can extend storage duration and improve the functional status of transplantable organs. This report characterizes the effect of SNO therapy on the corneal endothelial cells (CECs) during hypothermic storage, a key element in keratoplasty success.

Methods : Paired post-mortem human corneas (n=8) from research-consented donors were incubated in the hypothermic corneal storage solution (Life4^oC, Numedis, Inc.) with SNO agent or without SNO agent. CEC density and corneal thickness were measured at day 0, 7 and 14 using Konan CellChek D+ and OptoVue OCT machine, respectively. Total RNA from CEC layer was isolated using Qiagen MiniKit on day 14 and subjected to RNA-Seq to analyze differential gene expression pattern.

Results : Under normal circumstances during hypothermic storage, CEC density tends to decrease and corneal thickness tends to increase. CEC density was decreased by 8.2% for SNO without donors control group, while it remain unaltered for SNO group between day 7 and day 14. Corneal thickness was increased by 15% in the control group compared to only 8.6% increase in the SNO group between day 7 and 14. RNA-Seq analysis revealed more than 70 genes (transcripts) that were differentially expressed in the SNO group compared to the control group (P≤0.05, |Log₂Fold Change|≥0.5).

Conclusions : S-Nitrosylation therapy of hypothermic corneal storage might be beneficial to improve the quality of donated human corneas that are used for keratoplasty. With the significant alteration of transcriptome of the CEC with SNO therapy, these changes might contribute towards programming of the CECs to regulate long-term graft survival.

This is a 2021 ARVO Annual Meeting abstract.