Chrysin Attenuates Dry Eye Progression by Suppressing NOX2-Dependent Ferroptosis and STING/NLRP3-Mediated...

This study aimed to evaluate the effects of chrysin on dry eye and to elucidate its underlying mechanisms.

Public transcriptomic datasets were analyzed to identify ferroptosis-associated hub genes and to predict potential modulatory compounds. Ocular surface signs were evaluated using phenol red thread tests and corneal fluorescein staining. Chrysin was administered both in vivo and in vitro. Corneal ferroptosis and inflammatory levels were assessed by NOX2, 4-HNE, FACL4, GPX4, STING, and NLRP3 staining. HCE-T cells and primary mouse corneal epithelial cells were subjected to hyperosmotic conditions, whereas NOX2 and STING expression was modulated in HCE-T cells. Cell viability and mitochondrial morphology were evaluated using the CCK-8 assay and transmission electron microscopy. Intracellular levels of reactive oxygen species (ROS), Fe²⁺, and lipid peroxidation were measured using the fluorescent probes DCFH-DA, FeRhoNox-1, and BODIPY 581/591 C11, respectively.

There was a significant upregulation of NOX2 and STING under dry eye conditions. In hyperosmolarity-stimulated corneal epithelial cells, chrysin enhanced cell viability. In benzalkonium chloride-induced dry eye mice, chrysin restored tear volume and alleviated corneal epithelial damage. Chrysin suppressed the expression of NOX2, 4-HNE, and FACL4 while promoting GPX4 expression in the corneal epithelium. NOX2 overexpression reversed chrysin-mediated inhibition of ROS accumulation, Fe2+ overload, and lipid peroxidation in vitro. Furthermore, chrysin attenuated STING-dependent activation of the NLRP3 inflammasome and reduced IL-1β and IL-6 expression, which were abolished by STING overexpression.

Chrysin inhibits ferroptosis and inflammatory responses in corneal epithelial cells and improves ocular surface homeostasis in dry eye by suppressing the NOX2/ROS and STING/NLRP3 signaling pathways.