Abstract 1477: In Vitro and In Vivo Evidence for Hypoxia Inducible Factor 1-alpha Expression In Macrophages as Link between Vascular Inflammation and Angiogenesis
Background: Macrophages expressing the pro-angiogenic factor VEGF can contribute to plaque vulnerability via induction of neovascularization. Hypoxia Inducible Factor 1-alpha (HIF-1-alpha) has been identified as key transcriptional regulator of cellular VEGF-synthesis mediating the angiogenic tissue response to hypoxia. However, the role of HIF-1-alpha in macrophages in atherosclerosis is not well understood. Therefore, we assessed HIF-1-alpha and VEGF expression in peripheral blood-derived monocytes under normoxia and hypoxia with and without oxLDL in vitro and in macrophages of human atheroma in vivo.
Methods and Results: CD14+ mononuclear cells from peripheral blood of healthy human subjects were isolated using MACS cell sorting (purity >97%). Hif-1-alpha and VEGF were detected by immunofluorescence labeling. Under normoxia without oxLDL monocytes did not express HIF-1alpha and <10% stained positive for VEGF. Under normoxia and after 10 h incubation with ox-LDL (40Âμg/ml) 23Â±4% of cells expressed HIF-1alpha and 38Â±6% VEGF (P<0.05). Under hypoxia without oxLDL 28Â±5% and 40Â±8% of monocytes expressed HIF-1-alpha and VEGF, respectively (P<0.05. When combining hypoxia and ox-LDL up to 35Â±7% and 55Â±6% of monocytes expressed HIF-1-alpha and VEGF (P<0.05. Double-labeling demonstrated the coexpression of HIF-1-alpha and VEGF in single cells. In human atherosclerotic plaques (n=14) HIF-1-alpha and VEGF expression was evaluated by immunohistochemistry and morphometry. Both, HIF-1-alpha and VEGF were selectively increased in lipid-rich compared to fibrous plaque components (23Â±5 % vs. 5Â±2%, P<.01; 32Â±6% vs. 8Â±3%, P<.05, respectively) and correlated with macrophage content and degree of neovascularization. Double-labeling demonstrated colocalization of HIF-1-alpha and VEGF in macrophages close to neovessels.
Conclusions: The combined effect of ox-LDL and hypoxia on HIF-1-alpha and VEGF-expression in macrophages in vitro and the selective presence of HIF-1-alpha and VEGF in macrophages of lipid-rich plaque in vivo point to HIF-1-alpha as important link between inflammation and angiogenesis in vulnerable plaque.