Increased Vascular Endothelial Growth Factor 165 Binding to Kinase Insert Domain–Containing Receptor After Infection of Human Endothelial Cells by Recombinant Adenovirus Encoding the Vegf 165 Gene

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Abstract
Background —The angiogenic effect of vascular endothelial growth factor (VEGF 165 ) is mediated mainly through the high-affinity tyrosine kinase receptor VEGF-R2 (KDR/ flk-1 ). This study examined the effects of VEGF overexpression by primary human endothelial cells (ECs), which do not express VEGF under physiological conditions, on cell proliferation, VEGF binding to the kinase insert domain–containing receptor (KDR), and KDR expression. Methods and Results —Human primary ECs and SMCs were infected by recombinant adenoviral vector encoding VEGF 165 (rAdVEGF). Proliferation rate, bromodeoxyuridine incorporation, 125 I-labeled VEGF 165 binding to the KDR receptor, and KDR expression were tested in the infected cells and in cells supplemented with VEGF protein. Enhanced proliferation and a significant increase in 125 I-VEGF 165 binding to the KDR receptor were induced by rAdVEGF infection of ECs (autocrine effect) as well as by addition of recombinant VEGF 165 to noninfected cells. Infection of ECs by rAdVEGF led to posttranscriptional upregulation of the KDR receptor, whereas KDR mRNA expression levels remained unchanged. Similar effects were observed with supplemented recombinant VEGF 165 to noninfected ECs; nevertheless, this phenomenon occurred only with high VEGF 165 concentrations (10 ng/mL). Conclusions —The effect of VEGF 165 on proliferation and upregulation of KDR receptor expression demonstrated an autocrine phenomenon of EC sensitization. The fact that high concentrations of VEGF may be achieved in vivo by local continuous overexpression of VEGF 165 by gene transfer emphasizes the potential advantage of gene transfer over protein supplementation for therapeutic angiogenesis.