Vascular Endothelial Growth Factor Increases Human Cardiac Microvascular Endothelial Cell Permeability to Low-Density Lipoproteins

Background: Endothelial cell hyperpermeability is a proposed mechanism of increased lipid insudation into the vessel walls of allografts. Vascular endothelial growth factor (VEGF) is a potent inducer of vascular permeability and its expression is upregulated in human heart allografts. The goal of these experiments was to investigate the effects of VEGF on low-density lipoprotein (LDL) permeability through confluent monolayers of human cardiac microvascular endothelial cells (HCMEC) in vitro. Methods: VEGF mRNA and protein expression was characterized in coronary arteries from cardiac allograft vasculopathy patients as compared with healthy controls using in situ hybridization and immunohistochemical staining of sub-adjacent sections. HCMEC were grown to confluence and treated with VEGF-A₁₂₁ or VEGF-A₁₆₅. Permeability of LDL in confluent endothelial monolayers was measured using fluorometry. Transendothelial electrical resistance (TER) measurements were used to indirectly measure the tight junctional status. Immunocytochemical staining was performed to visualize changes in CD31 and zonula occludens-1. Results: We observed significant increases in VEGF expression within the superficial and deep intima and media of coronaries from allografts, as compared with controls. In vitro treatment with VEGF-A₁₂₁ and VEGF-A₁₆₅ significantly increased LDL passage through endothelial monolayers. We further showed that VEGF-A₁₂₁ and VEGF-A₁₆₅ caused significant decreases in TER at 2 to 4 hours post-treatment. Also, VEGF induced disruption of tight junctions, resulting in an increase in the intercellular gap formation. Conclusions: These results demonstrate that VEGF increases low-density lipoprotein permeability through endothelial monolayers, and this effect is correlated with VEGF-induced disruption of endothelial tight junctions resulting in the formation of intercellular gaps.