Abstract 490: Beta 3 Adrenoceptors Promote Proangiogenic Effects in Mouse and Human Endothelial Cells
β-adrenoceptors (βAR) play a key role, partly endothelium-dependent, in vascular homeostasis. We recently showed that endothelial β3AR mediate a sustained production of NO and vasodilation of human resistance arteries. Since NO is also a key mediator of angiogenesis, we assessed the effect of β3AR activation on angiogenesis and blood pressure (BP) regulation in vivo and in vitro. BP was measured by implanted telemetry in conscious mice. Compared to FVB controls, mice deficient in β3AR (β3KO) had higher SBP (124.1±0.6 vs 114.1±1.8, n=9 each, p<0.001), DBP (100.5±0.7 vs 95.41±1.6, n=9 each, p<0.01) than FVB controls and abolition of their circadian cycle, confirming the physiologic importance of β3AR in vivo. In a mouse aortic ring angiogenesis assay, 1nM of β3AR agonist SR58611A induced neocapillary tube formation (tubes/ring on day 12: 20.3±2.4 vs control 13.7±1.3, n=10 each, p<0.05). A replication-deficient adenovirus was generated to overexpress human β3AR in endothelial cells (Adβ3). β3AR overexpression induced phosphorylation of Protein Kinase B (PKB/Akt) in infected human microvascular endothelial cells (HMVEC). In subsequent angiogenesis assays infection of HMVECs on Matrigel-coated plates with Adβ3 resulted in a doubling of the length of capillary-like structures (99.1±10.6 μm) vs control AdGFP (47.4±17.5 μm, n=3, p<0.05). Likewise, using collagen gel-embedded spheroids of HUVECs, infection with Adβ3 enhanced the cumulative length of outgrowing sprouts (171.8±7.9 μm, n=7) vs control AdGFP (96.1±5.8 μm, n=4, p<0.01). The NO synthase inhibitor L-NAME (100μM) attenuated the effect (109.3±6.4 μm, n=6, p<0.05). Moreover, PI3 kinase inhibitor, LY294002 (15μM) significantly reduced the proangiogenic effect of β3AR adenovirus (81.9±12.9 μm, n=6, p<0.05). Finally, in a hindlimb ischemia model, cutaneous blood flow (normalized to controlateral limb, by laser Doppler) was also less at day 7 in β3KO (75.9±3%) vs FVB (92.5±3%, n=4 each, p<0.05) We conclude that the β3AR activates PKB/Akt pathway and mediates a NO-dependent proangiogenic effect in vitro and in vivo. Given the resistance of β3AR to homologous desensitization, this novel pathway may support a proangiogenic effect in response to ischemia with prolonged catecholamine stimulation.