Abstract 5278: The Sonic Hedgehog Transcription Factor Gli3 Modulates Ischemia-induced Angiogenesis
Gli transcription factors are mediators of hedgehog signaling and have been shown to be critical in several steps during development. We have shown that the Hedgehog pathway is reactivated in the adult cardiovascular system under ischemic conditions, however the specific role of Gli3 has not been elucidated. Adenoviral mediated overexpression of Gli3 promotes HUVEC migration (250±58% of control, p<0.001) while down regulation of Gli3 via siRNA delayed tube formation on Matrigel (total tube length after 8 hours 6.86 vs. 70.76 control), suggesting a possible role of Gli3 in angiogenesis. We next investigated the role of Gli3 in angiogenesis using Gli3+/− (Gli3+/XtJ) mice, a well established model of reduced Gli3 expression. VEGF-induced corneal angiogenesis was impaired in Gli3+/− mice compared to WT. The role of Gli3 in angiogenesis was then confirmed in two ischemia models. Hind-limb ischemia (HLI) was induced by resection of the left femoral artery. Capillary density was reduced by a mean of 48.40±12.08% in Gli3+/− mice vs. WT 7, 14 and 28 days. Myocardial infarction (MI) was induced by ligation of the LAD. 28 days after MI, left ventricular function assessed by echo and histological analysis revealed that Gli3+/− mice exhibit reduced ejection fraction (27.92±4.49% versus 37.56±7.02% for the WT, p=0.004), increased fibrosis area (33.65±9.73% versus 19.81±5.40% for the WT, p=0.007) and a decrease capillary density in the ischemic and border zones. These data indicate that Gli3 deficiency leads to impaired angiogenesis in both ischemic and non ischemic conditions. Moreover, the impairment in ischemia induced neovascularization is associated with more severe impairment of cardiac function after MI. The mechanism of Gli3’s effects was then investigated in vitro. Promoter reporter assays revealed that Gli3 overexpression inhibits Gli-dependent transcription, while Western analysis show increased Akt phosphorylation, activation of the ERK1/2 and increased c-Fos expression. Using a dominant negative Akt expressing virus and a MEK1/2 inhibitor, we show that Gli3 induced-EC migration is dependent on Akt and ERK1/2. These studies provide the first evidence that the Gli3 transcription factor regulates angiogenesis and EC phenotype.