Basic fibroblast growth factor induces expression of VEGF receptor KDR through a protein kinase C and p44/p42 mitogen-activated protein kinase-dependent pathway.

Abstract

Vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) are angiogenic molecules whose combined mitogenic activity is potently synergistic. However, the molecular mechanism underlying this synergy is incompletely understood. We examined whether VEGF and bFGF affect expression of each other or alter expression of the VEGF receptor KDR in retinal capillary endothelial cells. In addition, we investigated the intracellular signaling mechanisms involved in this response. VEGF-induced [3H]thymidine uptake was tightly correlated with KDR mRNA and protein concentrations, suggesting that increased KDR expression might account for VEGF's synergistic activity in the presence of bFGF. bFGF (10 ng/ml) induced KDR mRNA expression within 4 h and attained a 4.0-fold increase after 24 h. KDR protein expression was increased 7.5-fold after 48 h. VEGF (= 50 ng/ml) did not alter bFGF, VEGF, or KDR mRNA expression under serum-deprived conditions. In contrast, VEGF increased KDR mRNA expression 87% under growth conditions and 2.9-fold under serum-deprived conditions in the presence of bFGF. The protein kinase C (PKC) agonist phorbol myristate acetate (PMA) induced KDR mRNA expression 5.1-fold at 100 nmol/l. bFGF increased p44/p42 mitogen-activated protein kinase (MAPK) phosphorylation within 5 min, reaching a maximum within 15 min and remaining significantly elevated for >6 h. bFGF-induced MAPK phosphorylation and KDR mRNA expression were almost completely inhibited by 5 micromol/l GFX, a non-isoform-selective PKC inhibitor. MAPK inhibitor PD98059 reduced KDR mRNA expression 72% at concentrations that inhibited bFGF-induced MAPK phosphorylation 100%, suggesting that pathways in addition to MAPK might also be involved. Inhibitors of the beta isoform of PKC (LY333531), protein kinase A (PKA) (H89), and phosphotidylinositol (PI) 3 kinase (wortmannin) had no significant effect. These data suggest that bFGF stimulates KDR expression through a PKC and p44/p42 MAPK-dependent pathway not primarily involving the beta isoform of PKC, PKA, or PI-3 kinase. Since bFGF induces VEGF expression and since increased KDR expression potentiates VEGF action, resulting in additional KDR expression and marked mitogenic activity, these data provide a novel mechanistic explanation for the angiogenic synergy between VEGF and bFGF.

Vascular leakage in chick embryos after expression of a secreted binding protein for fibroblast growth factors

“…[37][38][39] This imbalance of vascular growth and differentiation signals was reported to lead to disruption of vascular integrity and to produce a phenotype reminiscent of that observed in the current study. 40,41 Our experiments comparing the effects of wild-type FGF-1 (without and with additional FGF-BP) and a form of FGF-1 engineered to be secreted supports the proposed mechanism of action of FGF-BP, that is, to enhance the activity of locally stored FGFs.…”

Fibroblast Growth Factor 2 Activation of Stromal Cell Vascular Endothelial Growth Factor Expression and Angiogenesis

“…sponse of endothelial cells to VEGF by upregulating the VEGFR-2 (flk-1/KDR) (Hata et al, 1999). Thus FGF-2-induced endothelial sensitivity to VEGF that is expressed within the developing stroma or secreted by tumor cells that are transformed could potentiate early tumor-associated stromal angiogenesis.…”

Two distinct signalling pathways are involved in FGF2-stimulated proliferation of choriocapillary endothelial cells: A comparative study with VEGF

“…It would be of value to test this possibility by investigating whether FGF2-induced CEC proliferation involves the autocrine action of VEGF. The induction of the expression of the VEGF receptor KDR by FGF2 in retinal capillary endothelial cells, con®rms that VEGF signalling is regulated by FGF2 (Hata et al, 1999). VEGF may play a central role in choroidal angiogenesis by mediating functions other than proliferation.…”

“…In order to rule out this hypothesis, it would be of value to analyse the e ects of FGF2 and VEGF on primary CEC cultures. It was recently reported that VEGF induced proliferation of macrovascular endothelial cells including human umbilical vein endothelial cells (HUVEC) and bovine aortic endothelial cells in vitro (Pedram et al, 1998;Yu and Sato 1999;, whereas it was une ective on bovine retinal capillary endothelial (BREC) cells in culture condition of low serum concentration (Hata et al, 1999). Overexpression of a negative dominant for FGFR1 in the choroid of mice induces a decrease in the normal neovascularization of the choroid during development (A Bikfalvi, personal communication).…”