Retinol-binding protein 3 in vitreous and plasma-derived small extracellular vesicles is reduced in proliferative diabetic retinopathy

Diabetic retinopathy (DR) is a severe microvascular complication of diabetes, progressing asymptomatically in its early stages and often leading to irreversible blindness. With the global diabetic population projected to reach 643 million by 2030, there is an urgent need for reliable predictive molecular signatures of vision-threatening DR (VTDR). Most reported circulatory biomarkers lack evidence of direct involvement in DR pathogenesis, underscoring the need for DR-specific factors that reflect retinal angiogenic pathophysiology. In this study, we utilized extracellular vesicles (EVs), lipid-encased nanovesicles known for their stability in biofluids, to explore the altered protein cargo of vitreous humor-derived small EVs (VH-SEVs) from patients with proliferative DR (PDR), an advanced stage of DR. Shotgun mass spectrometry identified retinol-binding protein 3 (RBP3), a photoreceptor-derived retinoid transporter with protective roles in DR, within VH-SEVs. VH-SEV-associated RBP3 levels were significantly reduced in PDR patients compared to those with macular hole (MH), as confirmed by immunoblotting and ELISA. Additionally, we detected RBP3 in plasma SEVs using immunoblotting and ELISA, revealing a decreasing trend in SEV-RBP3 levels across DR groups, with progressively lower levels in patients with non-proliferative DR (NPDR), and PDR. Notably, plasma SEV-RBP3 levels were significantly lower in diabetic patients with PDR compared to those without retinopathy. In conclusion, this study identifies RBP3, a DRrelevant retinal protein, within circulatory SEVs, highlighting its potential as a biomarker for VTDR and paving the way for its clinical applications.