Intermittent Hypoxia Destabilizes HIF-2

Abstract

Intermittent hypoxia, such as that occurring during transient repeated cessations in breathing for as little as 10 seconds in humans, elevates sympathetic nervous system activity and is associated with pathological conditions such as hypertension. Although continuous hypoxia increases the abundance of both members of the hypoxia-inducible factor (HIF) family (HIF-1α and HIF-2α), both of which dimerize with HIF-1β to form active transcription factors, Nanduri et al . show that these two isoforms respond oppositely to transient intermittent hypoxia. Cultured rat pheochromocytoma cells (PC12 cells) were exposed to 15, 30, or 60 cycles of intermittent hypoxia (30 seconds at 1.5% O 2 ) or to continuous hypoxia (4 hours at 1.5% O 2 ). Immunoblot analysis revealed that the abundance of HIF-1α increased under either intermittent or continuous hypoxia, whereas the abundance of HIF-2α decreased under conditions of intermittent hypoxia. The mRNA for HIF-2α actually increased in response to intermittent hypoxia, which suggests that protein stabilization was responsible. However, pharmacological inhibition of the proteasome or of the prolyl hydroxylases that are typically involved in controlling HIF stability had no effect on the decrease in HIF-2α that occurred in response to intermittent hypoxia. Instead, blocking an increase in intracellular calcium or pharmacological inhibition of calpain prevented the increase in HIF-2α triggered by intermittent hypoxia. The intermittent hypoxia-induced decrease in HIF-2α was associated with decreased activity of antioxidant enzymes, such as superoxide dismutase (SOD), and an increase in reactive oxygen species, indicative of oxidative stress. An SOD2 promoter reporter gene confirmed that intermittent hypoxia inhibited transcription, and this was overcome by forced expression of HIF-2α. To extend these findings in vivo, the authors treated rats with a calpain inhibitor and then exposed them to intermittent hypoxic conditions. In the carotid body and adrenal medulla of control rats exposed to intermittent hypoxia, the abundance of HIF-2α was decreased, as was SOD activity. These changes were prevented in the rats pretreated with the calpain inhibitor, and these rats also showed no elevation in blood pressure and no increase in circulating norepinephrine. Under conditions of intermittent hypoxia, calpain-mediated proteolysis appears to contribute to destabilization of HIF-2α, leading to oxidative stress and pathological changes in sympathetic tone. J. Nanduri, N. Wang, G. Yuan, S. A. Khan, D. Souvannakitti, Y.-J. Peng, G. K. Kumar, J. A. Garcia, N. R. Prabhakar, Intermittent hypoxia degrades HIF-2α via calpains resulting in oxidative stress: Implications for recurrent apnea-induced morbidities. Proc. Natl. Acad. Sci. U.S.A. 106 , 1199–1204 (2009). [Abstract][Full Text]