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Inhibition of Histone Methyltransferase G9a Activates Autophagy and Provides Protection in Dry Eye Disease

Year: 2025

Type: article

Abstract: Purpose: This study aimed to investigate the expression dynamics and functional role of the histone methyltransferase G9a in dry eye disease (DED). Methods: An in vitro DED model was established using human corneal epithelial cells (HCECs) cultured in hyperosmotic medium (90-mM NaCl). For the in vivo model, mice were treated with 0.075% benzalkonium chloride eye drops twice daily for 14 days. Phenotypic and mechanistic analyses were conducted using quantitative real-time PCR, western blotting, immunofluorescence staining, flow cytometry, and chromatin immunoprecipitation assays. Disease severity was assessed by fluorescein sodium staining and the phenol red thread test. Histopathological analysis included periodic acid-Schiff staining, hematoxylin and eosin staining, and immunohistochemistry. Results: G9a expression was significantly downregulated in the in vitro DED model, which was mediated by mammalian target of rapamycin (mTOR) signaling. Pharmacological inhibition of G9a mitigated DED-induced damage in HCECs by reducing inflammation, apoptosis, and oxidative stress while enhancing autophagic flux. Mechanistically, G9a inhibition alleviated transcriptional repression at the autophagy-related 7 (ATG7) promoter, decreasing H3K9me2 occupancy and promoting autophagy. In vivo, G9a inhibition improved disease outcomes, including (1) enhanced ocular surface integrity and increased tear production, (2) restoration of goblet cell density, and (3) reduced expression of corneal inflammatory markers. Conclusions: G9a downregulation confers protective effects in DED by promoting autophagy. Mechanistically, G9a inhibition enhances autophagy through the reduction of H3K9me2-mediated transcriptional silencing of ATG7. This G9a-autophagy axis offers significant protection against DED in both in vitro and in vivo models.

Source: Investigative Ophthalmology & Visual Science

DOI: 10.1167/iovs.66.14.52