Endoplasmic Reticulum Stress Induces ROS Production and Activates NLRP3 Inflammasome Via the PERK-CHOP Signaling Pathway in Dry Eye Disease.

Abstract

Purpose: The purpose of this study was to investigate the potential roles of endoplasmic reticulum (ER) stress in the development of dry eye disease (DED).

Methods: Single-cell RNA sequencing (scRNA-seq) data from the Gene Expression Omnibus (GEO) database, derived from corneal tissues of a dry eye mouse model, was processed using the Seurat R program. The results were validated using a scopolamine-induced dry eye mouse model and a hyperosmotic-induced cell model involving primary human corneal epithelial cells (HCECs) and immortalized human corneal epithelial (HCE-2) cells. The HCE-2 cells were treated with 4-phenylbutyric acid (4-PBA) or tunicamycin (TM) to modulate ER stress. TXNIP and PERK knockdown were performed by siRNA transfection. Immunofluorescence, Western blotting, and real-time PCR were used to assess oxidative stress, ER stress, unfolded protein response (UPR) marker proteins, and TXNIP/NLRP3 axis activation.

Results: The analysis of scRNAseq data shows an increase in the ER stress marker GRP78, and the activation of the PERK-CHOP of UPR in DED mouse. These findings were confirmed both in vivo and in vitro. Additionally, HCE-2 cells treated with 4-PBA or TM showed significant effects on the production of reactive oxygen species (ROS) and the activation of the TXNIP/NLRP3-IL1β signaling pathway. Furthermore, siRNA knockdown of PERK or TXNIP, which alleviated the TXNIP/NLRP3-IL1β signaling axis, showed protective effects on HCECs.

Conclusions: This study explores the role of ER stress-induced oxidative stress and NLRP3-IL-1β mediated inflammation in DED, and highlights the therapeutic potential of PERK-CHOP axis and TXNIP in the treatment of DED.