Vascular Endothelial Growth Factor-B Promotes In Vivo Angiogenesis

Abstract

Vascular endothelial growth factors (VEGFs) and their receptors have emerged as central regulators of the angiogenic process. However, involvement of VEGF-B, one of these factors, in angiogenesis remains obscure. Mice received subcutaneous injection of Matrigel alone or Matrigel with human recombinant protein rhVEGF-B 167 or with rhVEGF-A 165 . After 14 days, cell ingrowth in the Matrigel plug was increased by 2.0- and 2.5-fold in rhVEGF-B 167 –treated and rhVEGF-A 165 –treated mice, respectively ( P <0.01), in association with a raise in phospho-Akt/Akt (1.8-fold, P <0.01) and endothelial NO synthase (eNOS) (1.80- and 1.60-fold, respectively; P <0.05) protein levels measured by Western blot. VEGF-B–induced cell ingrowth was impaired by treatment with NOS inhibitor ( N G -nitro- l -arginine methyl ester; L-NAME, 10 mg/kg per day). Treatment with neutralizing antibody directed against the VEGF-B receptor VEGF-R1 (anti-VEGFR1, 10 μg) completely abrogated VEGF-B–related effects. Proangiogenic effect of VEGF-B was confirmed in a mouse model of surgically induced hindlimb ischemia. Plasmids containing human form of VEGF-A (phVEGF-A 165 ) or VEGF-B (phVEGF-B 167 or phVEGF-B 186 ) were administered by in vivo electrotransfer. Angiographic score at day 28 showed significant improvement in ischemic/nonischemic leg ratio by 1.4- and 1.5-fold in mice treated with phVEGF-B 167 and phVEGF-B 186 , respectively ( P <0.05). Laser Doppler perfusion data also evidenced a 1.5-fold increase in phVEGF-B 167 –treated and phVEGF-B 186 –treated mice ( P <0.05). Such an effect was associated with an upregulation of phospho-Akt/Akt and eNOS protein levels in the ischemic legs and was hampered by treatment with anti-VEGFR1. This study demonstrates for the first time that VEGF-B, in part through its receptor VEGF-R1, promotes angiogenesis in association with an activation of Akt and eNOS-related pathways.