Abstract
Purpose :
The blood-retinal barrier (BRB) mediates movement of molecules from the blood to the inner retina, protecting the retinal neural tissue from potentially harmful molecules and maintaining retinal homeostasis. Disruption of the BRB is associated with diabetic macular edema (DME). We and others have previously demonstrated that treatment with high doses of insulin is associated with increased incidence of BRB breakdown in diabetic mice and human type 2 diabetic patients. However, the molecular mechanisms involved remain poorly understood. The zebrafish is an advantageous model in which to investigate the molecular mechanisms of BRB disruption due to its small size, large clutches of embryos, genetic tractability, and similarity of their retinas to humans. We have previously demonstrated that retinoic acid (RA) signaling is necessary for maintenance of the zebrafish blood-retinal barrier. The purpose of this study was to determine whether treatment with high doses of insulin disrupts the BRB in zebrafish and whether RA signaling is involved in this BRB disruption.
Methods :
To visualize the BRB in vivo, we utilized the transgenic Tg(l-fabp:DBP-EGFP) zebrafish model that expresses vitamin D binding protein (a member of the albumin gene family) tagged to GFP. This model displays the integrity of the BRB with GFP-tagged protein localized within the retinal vasculature by 3 days post-fertilization. Breakdown of the BRB is visualized as “leaking” of GFP outside the vasculature. Zebrafish exposed to high levels of glucose in the presence or absence of insulin and/or all-trans retinoic acid (ATRA) were evaluated for BRB breakdown.
Results :
Zebrafish embryos exposed to glucose showed an increase in BRB disruption in the presence of insulin. Co-treatment with ATRA significantly attenuated the BRB disruption, suggesting that inhibition of RA signaling plays a role in insulin-mediated pathology. Treatment with insulin for 4 days resulted in a non-functional disrupted BRB in up to 95% of embryos. Adult zebrafish treated with insulin also showed BRB disruption.
Conclusions :
Here we establish a novel in vivo model of diabetes-induced BRB breakdown and demonstrate a potential role of RA signaling. In the future, this model can be used for high throughput drug screens and other studies to identify potential therapies for DME.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.