Abstract
Purpose :
Ischemic retinopathies (IR), the leading causes of vision loss, involve endothelial cell (EC) dysfunction, retinal neovascularization (RNV), and edema. Despite this, the mechanisms of blood-retinal barrier (BRB) disruption that leads to IR remain unclear. Our study revealed Akt activation and increased claudin-5 (Cldn5) expression in advanced glycation end-products (AGE)-treated human retinal endothelial cells (HRECs), and in a mouse model of oxygen-induced retinopathy (OIR). Interestingly, this correlated with a significant reduction in Claudin-17 (Cldn17) expression, an unexplored anion channel tight-junction protein. This research delves into the role of Cldn17 in HRECs and IR in mice.
Methods :
HRECs were used to investigate the impact of Cldn17 overexpression by adenovirus infection and knockdown using SiRNAs on cell migration, tube formation, and barrier resistance by electric cell-substrate impedance sensing (ECIS). Western blotting was performed to study the expression changes in Cldn5 and Cldn17. Newborn wild-type (WT) and Cldn17–/– (C57BL6/J) mice were subjected to OIR. Retinas were processed to study vascular damage using flat-mount analysis and immunofluorescence staining, and mice subjected to fundus fluorescein angiography (FA) to study retinal vascular permeability.
Results :
In our findings, we observed that overexpression of Cldn17 in HRECs led to a reduction in Cldn5 expression and an improvement in barrier resistance (n=4 and p<0.05). Conversely, when Cldn17 was knocked down, there was an increase in Cldn5 expression, accompanied by compromised barrier resistance (n=4 and p<0.05). Treatment with AGE and lipopolysaccharide (LPS) decreased the resistance of the HREC monolayer, but this effect was attenuated in HRECs overexpressing Cldn17 (n=6 and p<0.01). Additionally, AGE treatment facilitated HREC migration and tube formation, both of which were resisted by HRECs overexpressing Cldn17 (n=4, p<0.05). Confocal images of OIR retinal flat-mounts at postnatal day 17 stained with isolectin-B4, revealed increased vaso-obliteration and RNV in Cldn17–/– compared to WT (n=6 and p<0.01). Fluorescein angiography (FA) images demonstrated vascular hyper-permeability in Cldn17–/– OIR compared to WT OIR mice (n=6, p<0.01).
Conclusions :
Our findings underscore the significance of Cldn17 in maintaining the BRB integrity, and its depletion in IR is implicated in disease progression.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.