UP-REGULATION OF FLK-1 VASCULAR ENDOTHELIAL GROWTH-FACTOR RECEPTOR-2 BY ITS LIGAND IN A CEREBRAL SLICE CULTURE SYSTEM/

Vascular endothelial growth factor (VEGF) and its tyrosine kinase rece ptors VEGFR-1 (flt-1) and VEGFR-2 (flk-1/KDR) are key mediators of phy siological and pathological angiogenesis. They are expressed in most t issues during embryonic development but are down-regulated in the adul t, when angiogenesis ceases, Up-regulation of VEGFR-2 and of VEGF are observed in many pathological conditions under which angiogenesis is r einduced. A major regulator of VEGF expression is hypoxia. Although th e temporal expression pattern of VEGFR-2 parallels VEGF expression to a high extent, little is known about its regulation. Here, we show tha t VEGFR-2 is highly expressed in early postnatal mouse brain but is do wn-regulated commencing at postnatal day 15 (P15) of mouse brain devel opment and is hardly detectable in P30 mouse brain. Using P30 mouse br ain slices, we observed that hypoxia up-regulates VEGFR-2 in the slice s but not in human umbilical vein endothelial cells, suggesting the pr esence of a hypoxia-inducible factor in the murine neuroectoderm that up-regulates VEGFR-2. To identify the factors involved, normoxic P30 c erebral slices were cultured with growth factors that are either hypox ia-inducible (e.g., PDGF-BB, crythropoietin, and VEGF) and/or are know n to act on endothelial cells (e.g., PDGF-BB, VEGF, and P1GF). Exogeno usly added recombinant VEGF led to an up-regulation VEGFR-2 expression , which could be inhibited by preincubation with a neutralizing anti-V EGF antibody. Addition of PDGF-BB, P1GF, and erythropoietin had no eff ect on VEGFR-2 expression. Our results suggest a differential but syne rgistic regulation by hypoxia of VEGF and VEGFR-2: a direct induction of VEGF that subsequently up-regulates VEGFR-2 in endothelial cells. T his autoenhancing system mag represent an important mechanism of tumor angiogenesis.