Significance of Vascular Endothelial Cell Growth Factor Up-regulation Mediated Via a Chymase-angiotensin-dependent Pathway During Angiogenesis in Hamster Sponge Granulomas

Chymase is a serine protease responsible for local production of angiotensin (Ang) II from its precursor Ang I in several species, including humans, dogs, and hamsters. We have previously reported that chymase facilitates angiogenesis in sponge granulation tissues via local production of Ang II. Herein, we report the significance of vascular endothelial growth factor (VEGF) up-regulation mediated by Ang II during angiogenesis in hamster sponge granulomas. Treatment of granulation tissues with an anti-VEGF neutralizing antibody or antisense oligomers against VEGF mRNA significantly reduced Ang II-induced angiogenesis, supporting a significant role for VEGF during angiogenesis. In cultured fibroblasts prepared from granulation tissues, VEGF mRNA was up-regulated in response to Ang II within 2 h and this enhanced expression was abolished in the presence of an Ang II type 1 receptor-selective antagonist, an inhibitor of nuclear factor-kappaB activation, or an activator protein-1 inhibitor. To study the significance of local production of Ang II by chymase, we examined the effects of chymostatin on in vivo angiogenesis. We found that chymostatin markedly inhibited both up-regulation of VEGF mRNA and angiogenesis in granulation tissues treated by compound 48/80 or basic fibroblast growth factor. Our results suggest that Ang II directly acts on fibroblasts in granulation tissue to up-regulate VEGF mRNA and thereby induce angiogenesis. Furthermore, a chymase-Ang II-VEGF pathway may operate in granulation tissue as the primary mediator of angiogenesis.