Abstract 18269: Copper Transport Protein Antioxidant-1 is Involved in Post-Ischemic Neovascularization by Regulating Inflammatory Cell Recruitment and Angiogenesis
Neovascularization is important repair mechanism in response to ischemic injury and is dependent on angiogenesis and inflammation. Copper (Cu) plays an important role in regulating angiogenesis; however, its mechanism remains unknown. We previously demonstrated that Cu transport protein antioxidant-1 (Atox1) is involved in Cu-induced cell growth. We thus investigated the role of Atox1 in postnatal neovascularization using mouse hindlimb ischemia model. In vivo, Atox1 is expressed in endothelial cells (EC) and skeletal muscle basally, and following ischemia it is expressed in macrophages (mac3+). Atox1 knockout (KO) mice exhibit a marked reduction in post-ischemic blood flow recovery (BFR) at day 14, and capillary density (CD31+) and α-smooth muscle actin+ arterioles at day 7, compared to wild-type (WT) mice (57%, 45%, and 50% of WT, respectively). Furthermore, Atox1KO mice show decreased infiltration of macrophage into ischemic muscles at day 3 (20% of WT). Bone marrow (BM) transplantation reveals that Atox1 in tissue resident cells including ECs, but not BM derived cells, are required for ischemic neovascularization. Consistent with this, local injections of adenovirus driven Atox1 restores BFR in Atox1KO mice to that of WT. Mechanistically, adhesion molecules, ICAM1/VCAM1, mRNA levels in ischemic muscles are decreased in Atox1KO mice (15% and 5% of ischemic WT, respectively). In vitro, siAtox1 in human ECs blunted ICAM1/VCAM1 mRNA response to the TNFα (100U/ml 18h). Furthermore, siAtox1 reduces TNFα and VEGF induced phosphorylation of Akt (70% and 20% vs. stimulated con, respectively) in part due to loss of cytokine induced PTEN inactivation. Accordingly, these cells had increased susceptibility to apoptosis (DNA fragmentation). Moreover, VEGF-induced EC migration and tube formation are impaired by siAtox1. In conclusion, Atox1 plays an important role in regulating adhesion molecule gene expression as well as PTEN/pAkt pathway in ECs, thereby promoting inflammatory cell recruitment, EC survival, and angiogenesis in ischemic tissues, which may contribute to post-ischemic revascularization. These findings suggest Atox1 as a potential therapeutic target for inflammation and angiogenesis dependent ischemic cardiovascular diseases.
- © 2010 by American Heart Association, Inc.