Nitric oxide and reactive oxygen species exert opposing effects on the stability of hypoxia inducible factor-1α (HIF-1α) in explants of human pial arteries

Hypoxia induces angiogenesis, partly through stabilization of hypoxia-inducible factor-1alpha (HIF-1alpha), leading to transcription of pro-angiogenic factors. Here we examined the regulation of HIF-1alpha. by hypoxia and nitric oxide (NO) in explants of human cerebrovascular smooth muscle cells. Cells were treated with NO donors under normoxic or hypoxic (2% O-2) conditions, followed by analysis of HIF-1alpha. protein levels. Treatment with the NO donor sodium nitroprusside reduced levels of HIF-1alpha, whereas NO donors, NOC-18 and S-nitrosoglutathione, increased HIF-1alpha levels. SIN-1, which releases both NO and superoxide (O-2(.-)), reduced HIF-1alpha levels, suggesting that inhibitory NO donors may elicit effects through peroxynitrite (ONOO.-). O-2(.-) generation by xanthine/xanthine oxidase also reduced HIF-1alpha levels, confirming an inhibitory role for reactive oxygen species (ROS). Furthermore, superoxide dismutase increased HIF-1alpha levels, and the NO scavenger carboxy-PTIO reversed HIF-1alpha stabilization by NO donors. Effects on HIF-1alpha levels correlated with vascular endothelial growth factor transcription but did not affect HIF-1alpha transcription, as measured by RT-PCR and luciferase-reporter assays. The results indicate that HIF-1alpha is stabilized by agents that produce NO and reduce ROS but destabilized by agents that increase ROS, including O-2(.-) and ONOO.-. Thus we propose that the effect of NO on HIF-1alpha signaling is critically dependent on the form of NO and the physiological environment of the responding cell.