Investigating Late-Stage Diabetic Retinopathy: A Long-term Analysis of Vascular Changes in the Streptozotocin-Induced Mouse Model

Diabetic retinopathy (DR) is characterized by microvascular damage and increased vascular permeability in the retina. The investigation of visual outcomes in late-stage DR is limited by challenges of maintaining chronically hyperglycemic mice, and most reports are restricted to early-stage DR. In this study, we used carefully managed diabetic mice to longitudinally investigate associations between vascular leakage and visual acuity during early- and late-stage DR. Diabetes was induced in C57BL/6J mice with streptozotocin, and fluorescence angiography with dual fluorescence (FA-DF) was used to assess retinal vascular leakage dynamics in chronically hyperglycemic mice for 12 months. Retinal vascular leakage was evident 180 days after diabetes induction and before reduced visual acuity, measured using the optokinetic response, and vascular leakage continued to increase during DR progression. Mice were also treated with intravitreal injections of antiangiogenic aflibercept at late-stage DR, and reduced leakage was reliably measured using FA-DF and was associated with improved visual acuity. Inflammatory and vascular phenotypes were assessed using immunostaining, which revealed significantly lower retinal macrophage and vascular densities and reduced capillary diameter in association with anti-VEGF treatment compared with age-matched diabetic controls. In conclusion, this is the first longitudinal quantification of retinal vascular leakage in early, intermediate, and late stages of DR in the same cohort of mice in a minimally invasive fashion to demonstrate the associated effect of antiangiogenic therapy in vivo. Our findings also further confirmed the sensitivity of FA-DF in assessing retinal vascular leakage in conjunction with other functional measures in longitudinal studies in the same animals. ARTICLE HIGHLIGHTS We use the newly developed fluorescence angiography with dual fluorescence imaging method to longitudinally investigate associations between vascular leakage and visual acuity during early-, intermediate-, and late-stage diabetic retinopathy (DR) in diabetic mice. We demonstrate the onset and progression of vascular leakage, association of leakage with reduced visual acuity, and alteration of macrophage and vascular densities in late-stage DR. We confirm the sensitivity of fluorescence angiography with dual fluorescence in assessing retinal vascular leakage in conjunction with other functional measures in longitudinal studies in the same animals and demonstrate inflammatory changes in late-stage DR.