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Vascular endothelial growth factor (VEGF) is a newly identified growth and permeability factor with a unique specificity for endothelial cells. Recently the flt-encoded tyrosine kinase was characterized as a receptor for VEGF. A novel tyrosine kinase receptor encoded by the KDR gene was also found to bind VEGF with high affinity when expressed in CMT-3 cells. Screening for flt and KDR expression in a variety of species and tissue-derived endothelial cells demonstrates that flt is predominantly expressed in human placenta and human vascular endothelial cells. Placenta growth factor (PIGF), a growth factor significantly related to VEGF, is coexpressed with flt in placenta and human vascular endothelial cells. KDR is more widely distributed and expressed in all vessel-derived endothelial cells. These data demonstrate that cultured human endothelial cells isolated from different tissues express both VEGF receptors in relative high levels and, additionally, that all investigated nonhuman endothelial cells in culture are also positive for KDR gene expression.

Objective-Angiopoietin-2 (Ang-2) is a non-signal transducing ligand of the endothelial receptor tyrosine kinase Tie-2.Ang-2 is produced by endothelial cells and acts as an autocrine regulator mediating vascular destabilization by inhibiting Angiopoietin-1-mediated Tie-2 activation. To examine the transcriptional regulation of Ang-2, we studied the Ang-2 promoter in endothelial cells and nonendothelial cells. Methods and Results-The human Ang-2 promoter contains a 585-bp region around the transcriptional start site (Ϫ109 to ϩ476) that is sufficient to control endothelial cell-specific and cytokine-dependent Ang-2 expression. Strong repressor elements of Ang-2-promoter activity are located in the 5Ј-region of the promoter and in the first intron. The Ets family transcription factors Ets-1 and Elf-1 act as strong enhancers of endothelial cell Ang-2-promoter activity. Ets-binding sites Ϫ4 and Ϫ7 act as positive regulators, whereas Ets-binding site Ϫ3 acts as negative regulator. Demethylation experiments revealed that the Ang-2 gene (in contrast to the Tie-2 gene) is not controlled by imprinting. Key Words: angiogenesis Ⅲ Ang-2 Ⅲ Ets-1 Ⅲ Elf-1 Ⅲ endothelial cell A ngiogenesis, the formation of blood vessels from preexisting vessels, is controlled by a hierarchically structured signaling cascade of receptor tyrosine kinases specifically expressed in endothelial cells. 1,2 The angiopoietin-Tie ligand-receptor system plays a key critical role in the regulation of angiogenesis. The interactions of angiopoietins with the Tie-2 receptor regulate vascular maturation and vessel quiescence. 3 Angiopoietin-1 (Ang-1) consists of 4 alternatively spliced isoforms. 4 The 1.5-kb isoform codes for a multimerizing protein that acts in a paracrine manner and binds to endothelial cell-expressed Tie-2 inducing receptor phosphorylation and subsequent signal transduction. The structure of the smaller Ang-1 isoforms are consistent with dominant-negative regulatory molecules. 4 Ang-1-mediated activation of Tie-2 regulates endothelial cell survival and blood vessel maturation. Ang-1 exerts a vessel sealing effect, acts antiinflammatory, and protects against cardiac allograft arteriosclerosis. Low level constitutive Tie-2 activation may be required in the adult to maintain the mature quiescent phenotype of the resting vascular endothelium. 3 In contrast to the Tie-2 activating functions of Ang-1, Ang-2 acts primarily as functional antagonist of Ang-1/Tie-2 by binding the receptor without inducing signal transduction. 5,6 The opposing effects of Ang-1 and Ang-2 support a model of constitutive Ang-1/Tie-2 interactions controlling vascular homeostasis as default pathway 7 and Ang-2 acting as dynamically regulated antagonizing cytokine. 1,8,9 Loss of the Ang-2 gene and function is compatible with life as evidenced by the observation that Ang-2-deficient mice are born apparently normal. 6 The functionally unaffected blood vascular system of Ang-2-deficient mice has only minor abnormalities (eg, perturbed regression of hyaloid blood vessels). Yet ...

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