Tissue
hypoxia/
ischemia are major pathophysiological determinants. Conditions of decreased
oxygen availability provoke accumulation and activation of
hypoxia-inducible factor-1 (HIF-1). Recent reports demonstrate a crucial role of HIF-1 for inflammatory events. Regulation of hypoxic responses by the inflammatory mediators
nitric oxide (NO) and
reactive oxygen species (ROS) is believed to be of pathophysiolgical relevance. It is reported that hypoxic stabilization of HIF-1alpha can be antagonized by NO due to its ability to attenuate mitochondrial electron transport. Likely, the formation of ROS could contribute to this effect. As conflicting results emerged from several studies showing either decreased or increased ROS production during
hypoxia, we used experiments mimicking hypoxic intracellular ROS changes by using the redox cycling agent
2,3-dimethoxy-1,4-naphthoquinone (
DMNQ), which generates
superoxide inside cells. Treatment of A549, HEK293, HepG2, and COS cells with
DMNQ resulted in a concentration-dependent raise in ROS which correlated with HIF-1alpha accumulation. By using a HIF-1alpha-von Hippel-Lindau
tumor suppressor protein binding assay, we show that ROS produced by
DMNQ impaired
prolyl hydroxylase activity. When HIF-1alpha is stabilized by NO, low concentrations of
DMNQ (<1 microM) revealed no effect, intermediate concentrations of 1 to 40 microM
DMNQ attenuated HIF-1alpha accumulation and higher concentrations of
DMNQ promoted HIF-1alpha stability. Attenuation of NO-induced HIF-1alpha stability regulation by ROS was mediated by an active proteasomal degradation pathway. In conclusion, we propose that scavenging of NO by ROS and vice versa attenuate HIF-1alpha accumulation in a concentration-dependent manner. This is important to fully elucidate HIF-1alpha regulation under inflammatory conditions.