Vascular endothelial growth factor expression, β-catenin tyrosine phosphorylation, and endothelial proliferative behavior:: A pathway for transformation?

The hypothesis that tumor growth is angiogenesis dependent has been documented by a considerable body of direct and indirect experimental data and has generated intense basic and pharmaceutical-related interest. In contrast, the study of endothelial cell tumors has been modest by comparison. Hemangioma is the most common tumor of any kind seen in infancy and also, perhaps, the least understood. We compared a mouse hemangioma-derived cell line (EOMA) and primary human endothelial cells (HUVEC) for their proliferative behavior and molecular alterations. EOMA cells intrinsically expressed vascular endothelial growth factor (VEGF), which acts in an autocrine manner, resulting in an increase in CD1 expression and cell proliferation, both of which were inhibited by anti-VEGF neutralizing antibodies. Such an autocrine loop is supported by constitutive VEGF receptor (Flk-1) tyrosine phosphorylation, Flk-1 and FIt-1 nuclear localization, and mitogen-activated protein kinase activation. beta-catenin was also found to exhibit significant nuclear localization and constitutively associate with Flk-1 and FIt-1 in EOMA cells but much less so in HUVEC, and immunoprecipitated FIk-1 was able to phosphorylate purified P-catenin in an immune complex kinase assay. EOMA cells were also noted to express reduced levels of N-cadherin and gamma-catenin compared with HUVEC. Interestingly, sequestration of endogenous VEGF in EOMA cultures resulted in a dramatic decrease in nuclear M-catenin and a reduction in CD1 levels, whereas addition of exogenous VEGF elicited increased nuclear beta-catenin localization and increased CD1 levels in HUVEC. The possible contributions of VEGF signaling pathways, cell junction component expression levels, and phosphorylation states to endothelial cell transformation and proliferation are discussed.