Intermittent hypoxia has organ-specific effects on oxidative stress

Abstract

Obstructive sleep apnea is characterized by upper airway collapse, leading to intermittent hypoxia (IH). It has been postulated that IH-induced oxidative stress may contribute to several chronic diseases associated with obstructive sleep apnea. We hypothesize that IH induces systemic oxidative stress by upregulating NADPH oxidase, a superoxide-generating enzyme. NADPH oxidase is regulated by a cytosolic p47(phox) subunit, which becomes phosphorylated during enzyme activation. Male C57BL/6J mice were exposed to IH with an inspired O(2) fraction nadir of 5% 60 times/h during the 12-h light phase (9 AM-9 PM) for 1 or 4 wk. In the aorta and heart, IH did not affect lipid peroxidation [malondialdehyde (MDA) level], nitrotyrosine level, or p47(phox) expression and phosphorylation. In contrast, in the liver, exposure to IH for 1 wk resulted in a trend to an increase in MDA levels, whereas IH for 4 wk resulted in a 38% increase in MDA levels accompanied by upregulation of p47(phox) expression and phosphorylation. Administration of an NADPH oxidase inhibitor, apocynin, during IH exposure attenuated IH-induced increases in hepatic MDA. In p47(phox)-deficient mice, MDA levels were higher at baseline and, unexpectedly, decreased during IH. In conclusion, oxidative stress levels and pathways under IH conditions are organ and duration specific.

Fig. 1.

Expression of p47^phox protein in heart, aorta, and liver of mice exposed to 1 or 4 wk of intermittent air (control, IA) or intermittent hypoxia (IH). Each immunoblot represents results from 4 observations. In heart and aorta, IH did not induce changes in p47^phox expression or in serine phosphorylation (not shown) at 1 or 4 wk. In liver, IH induced an increase in p47^phox expression and serine phosphorylation after 4 wk. P-p47^phox, phosphorylated p47^phox.

Fig. 2.

Quantification by optical densitometry of immunoblots in Fig. 1. Open bars, IA; solid bars, IH. IH induced a 2-fold increase in hepatic p47^phox protein expression and phosphorylation. *P < 0.05 vs. IA.

Fig. 3.

Liver malondialdehyde (MDA) after exposure to 1 or 4 wk of IA or IH in mice treated with placebo or apocynin (2 mg·kg⁻¹·day⁻¹) or mice lacking functional p47^phox (p47^phox−/−). At 1 wk, there was a trend toward increased MDA with IH (P = 0.18). At 4 wk, IH induced a 38% increase in MDA. Apocynin attenuated the rise in MDA (*P < 0.005). In p47^phox−/− mice, increased MDA levels normalized during IH. Values are means ± SE; n = 96 (8 animals per group).

Fig. 4.

IH-to-IA MDA ratios in mice treated with placebo or apocynin (2 mg·kg⁻¹·day⁻¹) or p47^phox−/− mice exposed to 1 or 4 wk of IA or IH. At 4 wk, the time point at which IH resulted in significant increases in MDA, apocynin significantly attenuated the rise in liver MDA (*P < 0.01 vs. placebo). p47^phox−/− mice exhibited decreased MDA after IH as a result of increased oxidative stress during IA (†P < 0.005 vs. placebo). Decrease in MDA was greater in p47^phox than in apocynin-treated mice (P < 0.05). Error bars represent the sum of the standard errors of both IA and IH groups as a percentage of MDA total.