Fibroblast growth factor-2 selectively stimulates angiogenesis of small vessels in arterial tree.

Abstract

There is a critical need for quantifiable models of angiogenesis in vivo, and in general, differential effects of angiogenic regulators on vascular morphology have not been measured. Because the potent angiogenic stimulators fibroblast growth factor (FGF)-2 (basic FGF) and vascular endothelial growth factor (VEGF) are reported to stimulate angiogenesis through distinct signaling pathways, we hypothesized that FGF-2 stimulates vascular morphology differently than does VEGF and that stimulation of angiogenesis by FGF-2 is directly correlated to FGF receptor density. FGF-2 was applied at embryonic day 7 (E7), E8, or E9 to the quail chorioallantoic membrane (CAM); subsequent response of the arterial tree was measured by the fractal dimension (D(f)), a mathematical descriptor of complex spatial patterns, and by several generational branching parameters that included vessel length density (L(v)). After application of FGF-2 at E7, arterial density increased according to D(f) as a direct function of increasing FGF-2 concentration, and FGF-2 stimulated the growth of small vessels, but not of large vessels, according to L(v) and other branching parameters. For untreated control specimens at E7, L(v) of small vessels and D(f) were 11.1+/-1.6 cm(-1) and 1.38+/-0.01, respectively; at E8, after treatment with 5 microgram FGF-2/CAM for 24 hours, L(v) of small vessels and D(f) increased respectively to 22.8+/-0.7 cm(-1) and 1. 49+/-0.02 compared with 16.3+/-0.9 cm(-1) and 1.43+/-0.02 for PBS-treated control specimens. Application of FGF-2 at E8 and E9 did not significantly increase arterial density. By immunohistochemistry, the expression of 4 high-affinity tyrosine kinase FGF receptors was significantly expressed at E7, when CAM vasculature responded strongly to FGF-2 stimulation, but FGF receptor expression decreased throughout the CAM by E8, when vascular response to FGF-2 was negligible. In conclusion, the "fingerprint" vascular pattern elicited by FGF-2 was distinct from vascular patterns induced by other angiogenic regulators that included VEGF(165), transforming growth factor-beta1, and angiostatin.