Abstract 10: MicroRNA 17 in Angiogenesis: Lessons Learned From Immobilized Vascular Endothelial Growth Factor

Abstract

Several pre-clinical and clinical studies are exploring the therapeutic effect of cell-based therapies in ischemic diseases, including Peripheral Artery Disease (PAD). Unfortunately, most of the cells (more than 80%) die few days after delivery. We postulate that better understanding of VEGF Biology might be important for the design of more effective pro-survival strategies. Release of VEGF from ECM activates a carcinogenic program since it triggers vasculogenesis, tumor growth and metastasis. Contrarily, immobilized VEGF might have all the properties of soluble VEGF without inducing a carcinogenic program. Thus, identification of downstream players such as miRNAs mediating this process might be important. Herein, we evaluated therapeutic effect of miR-17 downregulation, which mediates the effect of immobilized VEGF both in vitro and in vivo. We recently showed that conjugated VEGF modulates cell activity by decreasing the expression miR-17 both in vitro and in vivo. In the present study, cell survival and angiogenesis were evaluated firstly in vitro using endothelial cells (ECs) transfected with antagomiR-17 to mimic the down-regulation of miR-17 by conjugated VEGF. AntagomiR-17 increased EC survival at least 1.5 times (n=6) compared to pro-angiogenic miRNAs reported in the literature (e.g. miR-424 and miR-132) and sprout formation on Matrigel at least 2 times (n=5) compared to all groups. The effect of antagomiR-17 was more pronounced under hypoxia conditions. In vivo, antagomiR-17 accelerated hemodynamic recovery of the whole limb (n=12) in unilateral limb ischemia obtained by occlusion of the left femoral artery. Blood flow recovery evaluated by Laser Doppler analysis was significantly higher 21 days after surgery in antagomiR-17 group compared to all other groups. Immunohistochemical analyses showed an increase in the capillary density of skeletal muscle in antagomiR-17 condition. In order to determine the gene target and potential pathway involved in the biological effect of antagomiR-17, next generation mRNA sequencing was performed. In conclusion, here we show the potential and underlying molecular mechanism of antagomiR-17 treatment in endothelial cell survival and angiogenesis both in vitro and in vivo.