The association of trimethylamine N-oxide with diabetic retinopathy Pathology: Insights from network toxicology and molecular docking analysis

Trimethylamine N-oxide (TMAO), a gut microbiota-derived metabolite, has emerged as a potential contributor to diabetic retinopathy (DR) progression. However, its molecular mechanisms in DR remain unclear. This study integrates network toxicology and multi-omics analyses to elucidate TMAO's role in DR pathogenesis. We identified TMAO-related targets through integration of CTD, SuperPred, and GeneCards databases. Differential expression analysis of DR-related genes was performed using GSE60436 and GSE102485 datasets. We intersected these with TMAO targets to identify key genes. Functional enrichment and pathway analyses were conducted, followed by immune cell infiltration assessment using ssGSEA. Machine learning algorithms (LASSO and RF) identified key marker genes, validated through GSE94019 dataset and in vitro experiments. Molecular docking explored interactions between TMAO and key proteins. We identified 45 TMAO-related targets implicated in DR. Functional analysis revealed enrichment in stress response and inflammatory pathways. Differential pathway analysis indicated significant upregulation of immune and apoptotic pathways in DR. Immune cell infiltration analysis showed increased levels of cytotoxic and inflammatory cells in DR. CASP3, CXCR4, and MAPK1 emerged as key marker genes, their expression significantly upregulated in PDR patients. Molecular docking highlighted stable interactions between TMAO and these proteins, suggesting potential modulation of their activity. TMAO-associated targets are enriched in inflammatory, oxidative, and apoptotic pathways in PDR tissues, suggesting a potential (but not causal) link to DR pathology. Our findings highlight the gut-retina axis in DR and provide a framework for targeting TMAO-mediated mechanisms in diabetic complications.