A Novel Methodology to Assess Retinal Vascular Permeability in a Rodent Model of Diabetes

Abstract

PurposeDiabetic retinopathy (DR) is a major complication of diabetes and is one of the leading causes of blindness worldwide. Treatment of DR can only be achieved through an enhanced understanding of disease pathogenesis. Structural, functional and biochemical studies cannot be carried out in human subjects and chemically induced diabetic rodent models that mimic the early developmental changes in DR and have become the gold standard. In combination with fundus fluorescein angiography (FFA) retinal abnormalities, non‐perfusion and vascular permeability can be assessed and in response to treatments over time.MethodsFFA was carried out in non‐diabetic and diabetic Norway Brown rats (250–300g) on days 0, 14, 21 and 28 using the Micron IV retinal imaging microscope (Phoenix Research Labs, CA). Animals received a single dose of PBS (n=6) or streptozotocin (STZ) i.p., 50mg/kg (n=6) and blood glucose levels were measured throughout. Video angiograms were quantitatively analysed for suboptical resolution microvascular permeability by using the arterioles as the measure for luminal concentration. In a separate subset of experiments primary human retinal endothelial cells (pHREC) were exposed to normo‐(D‐Mannitol, 30mM) or hyperglycaemia (D‐Glucose, 30mM) and probed for glycosaminoglycans (GAGs) heparan sulfate (HS), chondroitin sulfate (CS), and the tight junction marker ZO‐1.ResultsDiabetes was successfully induced in Norway Brown rats following a single dose of STZ (Blood glucose 28.4±2.6 mmol/L, day 28). By adapting Fick's Law retinal permeability (Ps, um.s^−1) was calculated and shown to significantly increase (p<0.05) for each time point, over the 28‐day period. To understand the mechanisms that may account for this increase in solute permeability across the retinal vessel wall we observed a significant decrease in sulphated GAG expression, more specifically HS (p<0.05) in pHREC exposed to hyperglycaemia versus normoglycaemia. In addition, the expression of the tight junction protein ZO‐1 significantly decreased in pHREC when exposed to hyperglycaemic conditions.ConclusionsRetinal vascular solute permeability can be accurately quantified in a rodent model of diabetes using the outlined methodology and mathematical quantification. Advanced quantitative assessments such as this could substantially contribute to a better understanding of the pathogenesis of DR and identify potential drugs for treatment.Support or Funding InformationMRCThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.