Aqueous Humor of Failed Corneal Grafts Induce Inflammasome Activation in Endothelial Cells

Abstract

Purpose : High levels of proinflammatory cytokines in the aqueous humor (AqH) have been associated with corneal graft failure (CGF) and endothelial cell loss after corneal transplant. Pyroptosis is a recently described caspase-1-dependent proinflammatory programed cell death mechanism. Pyroptosis is triggered upon inflammasome activation, which is a multiprotein complex comprised by the protein apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC); a nucleotide oligomerization domain-like receptor (NLRs); and procaspase-1. In the present study, we assessed caspase-1 levels in primary human corneal endothelial cells (PhCEC) following exposure to the AqH of patients with CGF and from healthy patients.

Methods : PhCEC were incubated in vitro for 60 minutes with the AqH of patients with CGF (n=4) who underwent corneal transplantation and healthy patients who underwent cataract surgery (n=4). The levels of caspase-1 in PhCEC were determined by ELISA 24 hours following exposure. A specific caspase-1 inhibitor (Ac-YVAD-cmk) was used to assess downregulation in these cells.

Results : Protein levels of caspase-1 were significantly higher in PhCEC incubated with the AqH of patients with CGF than those incubated with the AqH of healthy eyes (p<0.0043), and with the group with no AqH exposure (p<0.0008). The last two groups did not show differences between them (p=0.58). Additionally, levels of caspase-1 were significantly downregulated by the caspase-1 inhibitor Ac-YVAD-cmk in PhCEC exposed to the AqH of patients with CGF (p<0.0017) but showed no differences in other groups.

Conclusions : AqH of patients with CGF induced inflammasome activation in corneal endothelial cells and was downregulated by Ac-YVAD-cmk, suggesting that pyroptosis may play an essential role in the pathogenesis of endothelial cell death that leads to CGF. These findings reassert the importance of further studies of this pathway for therapeutic purposes.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.