Integrated metabolomics and transcriptomics reveal retinal alterations in a mouse model of chronic restraint stress protocol-induced anxiety-like behavior

Background Psychological stress is a potential risk factor for glaucoma, but the association and underlying molecular mechanisms remain unclear. This study investigated whether a chronic restraint stress (CRS) protocol, which includes repeated fasting/water deprivation, is associated with glaucoma-like retinal damage in mice and explored the involved pathways. Methods Adult male mice underwent 4 h daily restraint for 10 days. Anxiety-like behaviors were assessed by open field test (OFT) and elevated plus maze (EPM). Intraocular pressure (IOP) was measured at baseline, day 7, and day 14. Retinal ganglion cell (RGC) survival was quantified by RBPMS immunofluorescence. RNA-seq and untargeted metabolomics (UHPLC-MS/MS) were performed on retinas, followed by integrated analysis, qPCR validation, and a targeted query of the GWAS Catalog for glaucoma-associated SNPs. Results CRS mice showed anxiety-like behaviors, progressive IOP elevation, and reduced RGC numbers. Transcriptomics identified 216 candidate genes with nominal differential expression, with altered oxidoreductase activity and iron ion binding. Metabolomics detected 255 differential metabolites, mainly fatty acids, glycerophospholipids, and amino acids, with nominal enrichment of the thermogenesis pathway. qPCR confirmed upregulation of Cyp2f2, Cyp4f15, Fmo2, S100a9, and Aif1. Integrated analysis identified Fmo2 and Serpine1 as candidate genes that may contribute to the link between the CRS protocol and glaucoma pathogenesis; Serpine1 had records of glaucoma-risk SNPs in the GWAS Catalog. Conclusion The CRS protocol is associated with glaucoma-like retinal degeneration, characterized by IOP elevation, RGC loss, and dysregulation of metabolic, oxidative stress, and inflammatory pathways. Fmo2 and Serpine1 are candidate genes that may link the CRS protocol to glaucoma pathogenesis; however, functional validation is required to establish causality.