A novel co‐culture model of the blood‐retinal barrier based on primary retinal endothelial cells, pericytes and astrocytes

Abstract

AbstractPurpose Loss of blood‐retinal barrier (BRB) is an important cause of diabetic macular edema (DME), but cellular mechanisms underlying BRB dysfunction are poorly understood. Therefore, we developed and characterized a novel in vitro BRB model.Methods The model is based on primary bovine retinal endothelial cells(BRECs).These cells were shown to maintain specific in vivo BRB properties by expressing high levels of endothelial junction proteins and specific BRB transporters. To investigate the influence of pericytes and astrocytes on BRB maintenance in vitro, we compared five different co‐culture BRB models, based on BRECs, bovine retinal pericytes (BRPCs) and rat glial cells.Results Co‐cultures of BRECs with BRPCs and glial cells showed the highest trans‐endothelial resistance (TEER) as well as decreased permeability of tracers, even after vascular endothelial growth factor (VEGF) stimulation, suggesting a major role for these cell types in maintaining barrier properties. To mimic the in vivo situation of DME, we stimulated BRECs with VEGF, which downregulated MDR1 and GLUT1 mRNA levels, transiently reduced expression levels of endothelial junctional proteins and altered their organization, increased the number of intercellular gaps in BRECs monolayers and influence the permeability of the model to differently‐sized molecular tracers. Moreover, as has been shown in vivo, expression of plasmalemma vesicle‐associated protein (PLVAP) was increased in endothelial cells in the presence of VEGF.Conclusion This in vitro model is the first co‐culture model of the BRB that mimicks in vivo VEGF‐dependent changes occurring in DME.