Differential effect of inflammatory mediators on the inner and outer blood-retinal barrier

Abstract

Purpose : The integrity of the blood-retinal barrier (BRB) represents an essential prerequisite for retinal function and visual acuity. It is a complex structure consisting of an outer unit (oBRB) formed by the lateral tight junctions of the retinal pigment epithelium (RPE) and an inner unit (iBRB) formed by vascular endothelium. Inflammatory stimuli in the context of retinal disease like diabetic retinopathy (DR) and age-related macular degeneration (AMD) can lead to loss of function of the BRB with oedema formation and accumulation of lipid and protein deposits. In this study, we compare the influence of inflammatory lipid (prostaglandins) and cytokine (Interleukin 6, IL6) mediators relevant to AMD and DR on the iBRB and oBRB in vitro.

Methods : Real-time cell analysis (RTCA) impedance measurements were performed using human umbilical vein endothelial cells (HUVEC) and retinal pigment epithelial cells (ARPE-19). Prostaglandin E2 (PGE2), Prostaglandin F2α (PGF2α) and IL6 with its soluble receptor (sIL6R) were used to stimulate both cell lines. The expression and localisation of zonula occludens (ZO-1) was characterised using immunocytochemistry.

Results : A single treatment of ARPE-19 cells with PGE2 led to a significant short-time reduction in impedance compared to carrier-treated cells (p<0.05), while PGF2α had no effect. Immunocytochemical staining for ZO-1 in ARPE-19 cells treated with PGE2 showed a serrated morphology and reduction of ZO-1 expressing cell-cell junctions compared to controls. Interestingly, prostaglandins did not have any effect on HUVECs in impedance measurements as well as ZO-1 staining patterns.
In contrast to prostaglandins, IL6 + sIL6R significantly decreased impedance in RTCA measurements (p<0.05) in ARPE-19 cells as well as HUVECs with delayed onset and prolonged duration. These functional changes correlated with significant reduction in expression (ARPE-19, HUVEC) as well as serrated morphology of ZO-1-bearing cell junctions in ARPE-19 cells.

Conclusions : Inflammatory mediators induce the breakdown of the BRB in a stimuli- and cell-type specific manner. In our cell culture model, PGE2 mainly affects the oBRB suggesting its potential influence in AMD while IL6 + sIL6R causes distinct changes in the iBRB and oBRB. However, the kinetics of PGE2 and IL6 + sIL6R differ indicating distinct mechanisms in the oBRB breakdown.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.