Regulation of the hypoxia-inducible factor 1α by the inflammatory mediators nitric oxide and tumor necrosis factor-α in contrast to desferroxamine and phenylarsine oxide

Hypoxic/ischemic conditions provoke activation of the hypoxia-inducible factor-1 (HIF-1), which functions as a transcription factor. HIF-1 is composed of the HIF-1 alpha and -beta subunits, and stability regulation occurs via accumulation/degradation of HIF-1 a with the notion that a prolyl hydroxylase accounts for changes in protein level. In addition, there is evidence that HIF-1 is up-regulated by diverse agonists during normoxia. We investigated the impact of inflammatory mediators nitric oxide (NO) and tumor necrosis factor-alpha (TNF-alpha) on HIF-1 alpha regulation. For comparison, LLC-PK1 cells were exposed to hypoxia, stimulated with desferroxamine (DFX, known to mimic hypoxia), and the thiol-crosslinking agent phenylarsine oxide (PAO). Although all stimuli elicited HIF-1 alpha stabilization with differences in the time-dependent accumulation pattern, significant variations appeared with regard to signaling. With the use of a superoxide anion (O-2(-)) generator, we established an O-2(-)-sensitive pathway that blocked HIF-1 alpha stabilization in response to NO and TNF-alpha while DFX- and PAO-evoked HIF-1 alpha stabilization appeared O-2(-)-insensitive. NO and TNF-alpha signaling required phosphorylation events, especially activation of the phosphatidylinositol 3-kinase/Akt, which is in contrast to DFX and PAO. Based on HIF-1-dependent luciferase reporter gene analysis, it was found that, in contrast to NO and TNF-alpha, PAO resembled a stimulus that induced a dysfunctional HIF-1 complex. These data indicate that diverse agonists activate HIF-1 a under normoxic conditions by employing different signaling pathways.