ID3 inhibits the HSV-1 replication through the ROS/ ID3/Wnt signaling axis in the pathogenesis of herpes simplex keratitis

Herpes simplex virus type 1 (HSV-1) is a widespread pathogen affecting a large global population. HSV-1 infection can induce herpes simplex keratitis (HSK), a leading cause of vision loss. Current therapeutic approaches remain limited in efficacy. Consequently, investigating novel pathogenic mechanisms is essential for developing improved treatment strategies. Whether reactive oxygen species (ROS) regulates HSV-1 infection in corneal epithelial cells (CECs) through the DNA-binding protein inhibitor ID3 (ID3), and the underlying mechanism, has not been previously reported. This study demonstrated that HSV-1 infection elevates ROS levels and apoptosis in both human CECs and HSK mouse models. The antioxidant N-acetylcysteine (NAC) reversed these effects. Whole-transcriptome sequencing, RT-qPCR, and immunofluorescence analyses revealed that ID3 expression increased in NAC-pretreated human CECs and HSK mouse models. Overexpression of ID3 suppressed ROS accumulation, HSV-1 replication, and apoptosis in infected CECs, as confirmed by western blotting, flow cytometry, immunofluorescence, and in-cell western assays. Furthermore, ID3 overexpression downregulated the Wnt/β-catenin signaling pathway, which correlated with reduced HSV-1 replication and diminished apoptosis in CECs. In HSV-1-infected CECs, elevated ROS promotesHSV-1 replication and apoptosis by inhibiting ID3 expression, thereby activating the Wnt/β-catenin signaling pathway. These findings identify ID3 as a promising therapeutic target for HSK.