Sonic hedgehog induces angiogenesis via Rho kinase-dependent signaling in endothelial cells

Abstract

The morphogen Sonic Hedgehog (Shh) promotes neovascularization in adults by inducing proangiogenic cytokine expression in fibroblasts; however, the direct effects of Shh on endothelial cell (EC) function during angiogenesis are unknown. Our findings indicate that Shh promotes capillary morphogenesis (tube length on Matrigel ™ increased to 271±50% of the length in untreated cells, p=0.00003), induces EC migration (modified Boyden chamber assay, 191±35% of migration in untreated cells, p=0.00009), and increases EC expression of matrix metalloproteinase 9 (MMP-9) and osteopontin (OPN) mRNA (real-time RT-PCR), which are essential for Shhinduced angiogenesis both in vitro and in vivo. Shh activity in ECs is mediated by Rho, rather than through the "classic" Shh signaling pathway, which involves the Gli transcription factors. The Rho dependence of Shh-induced EC angiogenic activity was documented both in vitro, with dominantnegative RhoA and Rho kinase (ROCK) constructs, and in vivo, with the ROCK inhibitor Y27632 in the mouse corneal angiogenesis model. Finally, experiments performed in MMP-9-and OPNknockout mice confirmed the roles of the ROCK downstream targets MMP-9 and OPN in Shhinduced angiogenesis. Collectively, our results identify a "non-classical" pathway by which Shh directly modulates EC phenotype and angiogenic activity.

Dichotomy in Hedgehog Signaling between Human Healthy Vessel and Atherosclerotic Plaques

“…The in vivo relevance of the induction of these genes was proven in ischemia experiments, in which intramyocardial gene transfer of SHH promoted recovery and preservation of left ventricular function in myocardial ischemia by enhanced neovascularization (11), supporting the notion that HH aids in ischemic tissue rescue and especially in its revascularization. In agreement, HH production is critical for VEGF production (9,12) and endothelial maintenance and growth (9,(13)(14)(15)(16)(17). From this, we can infer that HH signaling may well play a detrimental role as well, causing revascularization-mediated plaque instability, and constitutes an attractive candidate for targeting pathological neovascularization (18).…”

Hedgehog-Dependent Regulation of Angiogenesis and Myogenesis Is Impaired in Aged Mice

“…This latter result is consistent with a previous study demonstrating that Shh-induced angiogenesis in the mouse cornea is independent of Gli1. 29 Together these data suggest that Hh-dependent regulation of angiogenesis and myogenesis involves distinct mechanisms. Moreover, these data imply that restoring nerve density or promoting angiogenesis is not sufficient to promote myogenesis at least in old animals.…”

Autonomous and nonautonomous roles of Hedgehog signaling in regulating limb muscle formation

“…In cell culture studies, Shh has been shown to be required for cell motility by activating Rho GTPases through a ''noncanonical'' or Gli-independent pathway (Renault et al 2010;Sasaki et al 2010;Polizio et al 2011). In these scenarios, signaling activation is relayed from Smo to the G proteins or Tiam1, a GEF for Rac1 (Sasaki et al 2010;Polizio et al 2011).…”