Diabetes mellitus is associated with an increased risk for cardiovascular diseases. The relationship between hyperglycemia, endothelial dysfunction and vascular disorders is well established. However, the precise mechanism of the initiation and progression of vascular lesions in diabetic patients remains still unclear. The transcription factor COUP-TFII regulates transcriptional activity of the rat insulin promoter. A heterozygous deletion in mouse islet ß-cells leads to impaired glucose sensitivity and abnormal insulin secretion. Furthermore, COUP-TFII is specifically expressed in venous endothelium and has an influence on arteriovenous differentiation. Therefore, we cultured venous endothelial cells (HUVEC) in high glucose medium and stimulated them with 5nM insulin at different time points for up to 48 h. First, we verified expression of glucose transporters GLUT1, 3, 6 and 10 and insulin receptor in cultured primary human endothelial cells. After stimulation with 5nM insulin we found a strong and significant increase of Akt phosphorylation on Ser473, which is impended by PI3 kinase inhibition. After stimulation for up to 60min, we found a significant increase of COUP-TFII expression with or without insulin in HUVEC. In contrast, long term culture in high glucose medium with or without insulin stimulation resulted in a significant decrease of COUP-TFII protein, even while COUP-TFII mRNA is not significant regulated. E-selectin and insulin receptor expression were strongly upregulated after long-term stimulation with insulin. In conclusion, our data suggest a regulation of COUP-TFII by glucose in venous endothelial cells.