Proteomic Analysis Reveals Dysregulated Tear Fluid Proteins in Glucose Metabolism and HIF-1 Signaling in Diabetes

Purpose : Diabetes is the most prevalent metabolic disorder worldwide, and research into diabetic eye complications has shown potential of using tear fluid as a source of non-invasive biomarkers. However, the relationship between the tear fluid proteome and diabetes remains unclear. We hypothesize that diabetes dysregulates glycolytic proteins and other risk factors associated with diabetic eye complications.

Methods : Tear fluid was collected using Schirmer's strips from age-matched participants (58.8 ± 5 yrs old, p=0.43) with type 2 diabetes mellitus (DM, n=10) and healthy controls (HC, n=10). Fasting blood glucose levels were significantly higher in the diabetes group (9.1±2.7 mmol/L) compared to healthy controls (5.8±0.4 mmol/L, p=0.003). Tear fluids were prepared using S-Trap sample preparation as previously described. Proteins (2 µg) were quantified using SWATH-MS acquisition on a Sciex TripleTOF 6600 mass spectrometer. Data processing was performed with library-free quantification utilizing DIA-NN software. Statistical analysis was performed using a two-tailed Student's t-test.

Results : Tear fluid analysis identified a total of 2652 protein groups at a 1% false discovery rate (FDR). There were 33 dysregulated proteins with a fold-change (DM/HC) of ≥1.3 or ≤0.77, and significance p<0.05. Gene ontology analysis revealed differentially expressed proteins involved in pyruvate metabolism (FDR = 0.0002). Additionally, KEGG functional enrichment analysis revealed significant associations with pathways related to glycolysis or gluconeogenesis (FDR = 2.40E-07), and the HIF-1 signaling pathway (FDR = 0.002). These findings suggest that metabolic dysregulation and hypoxic responses are prominent in the tear fluid of diabetic patients.

Conclusions : Our results identified dysregulated proteins associated with pyruvate metabolism, glycolysis, and HIF-1 signaling pathway in diabetes. These findings reinforce the connection between tear fluid and diabetes, emphasizing the influence of glycemic control on tear fluid composition and the role of HIF-1 signaling in the pathogenesis of diabetic complications. The identified proteins hold promise as potential biomarkers for screening diabetes and its associated ocular complications, providing insights for further research.

This abstract was presented at the 2025 ARVO Annual Meeting, held in Salt Lake City, Utah, May 4-8, 2025.