Abstract 16029: Activator of G-Protein Signaling 8, an Ischemia-Inducible G-Protein Activator, is Required for Vascular Endothelial Growth Factor-Mediated Collateral Development
Ischemia elicits multiple events in the heart, including myocardial cell death and coronary collateral development. These events are mediated by G-protein coupled receptors, but are also regulated by receptor-independent activators of heterotrimeric G-proteins. We previously identified one such regulatory protein, activator of G-protein signaling 8 (AGS8), from a rat heart with substantial collaterals following repetitive transient ischemia. AGS8 is up-regulated in response to hypoxia/ischemia and directly interacts with Gβγ. AGS8 plays a pivotal role in the hypoxia-induced apoptosis of cardiomyocytes by regulating Gβγ signaling. However, the role of AGS8 in collateral development has not previously been investigated. Here, we examine the role of AGS8 in endothelial cells by siRNA-mediated knockdown of AGS8. AGS8-siRNA inhibited HUVEC tube formation on fetal calf serum-stimulated matrigel (23.3 ± 2.9% of control, p<0.01, mean±SEM). Vascular endothelial growth factor (VEGF)-induced endothelial cell tube formation was also attenuated by AGS8-siRNA (27.0 ± 4.8% of control, p<0.01, mean±SEM), as was VEGF-stimulated cell growth (MTT assay: 30.5 ± 3.7% of control, p<0.01, mean±SEM). VEGF (25 ng/ml) stimulated phosphorylation of p42/44 MAPK and p38 MAPK; however, knockdown of AGS8 inhibited these signaling events (phospho-p42/44MAPK / p42/44MAPK: 29.5 ± 9.5% of control, p<0.05. phospho-p38MAPK / p38MAPK: 36.2 ± 6.1% of control, p<0.05. mean±SEM). Interestingly, phosphorylation of VEGF receptor-2 (KDR) by VEGF was completely blocked by AGS8-siRNA (phospho-KDR (Tyr996) / KDR: 3.4 ± 1.4% of control, p<0.01. phospho-KDR (Tyr1175) / KDR: 2.1 ± 1.6% of control, p<0.01. mean±SEM). Moreover, the AGS8-peptide, designed to disrupt AGS8-Gβγ interaction, successfully blocked the association of Gβγ with AGS8 and inhibited VEGF-induced tube formation. These data suggest that AGS8-Gβγ interaction is required for VEGF signaling linked to myocardial collateral development, and that AGS8 plays a key role in myocardial adaptation to hypoxia in cardiomyocytes and endothelial cells.
Author Disclosures: H. Hayashi: None. A.A. Mamun: None. M. Sakima: None. H. Suzuki: None. M. Sato: None.
- © 2014 by American Heart Association, Inc.