Chronic hypoxia impairs cytochrome oxidase activity via oxidative stress in selected fetal Guinea pig organs

Abstract

We hypothesized that chronic hypoxia disrupts mitochondrial function via oxidative stress in fetal organs. Pregnant guinea pig sows were exposed to either normoxia or hypoxia (10.5% O2, 14 days) in the presence or absence of the antioxidant, N-acetylcysteine (NAC). Near-term anesthetized fetuses were delivered via hysterotomy, and fetal livers, hearts, lungs, and forebrains harvested. We quantified the effects of chronic hypoxia on cytochrome oxidase (CCO) activity and 2 factors known to regulate CCO activity: malondialdehyde (MDA) and CCO subunit 4 (COX4). Hypoxia increased the MDA levels in fetal liver, heart, and lung with a corresponding reduction in CCO activity, prevented by prenatal NAC. The COX4 expression paralleled CCO activity in fetal liver and lung, but was unaltered in fetal hearts due to hypoxia. Hypoxia reduced the brain COX4 expression despite having no effect on CCO activity. This study identifies the mitochondrion as an important target site in tissue-specific oxidative stress for the induction of fetal hypoxic injury.

Figure 1.

Fetal guinea pig body and organ weights. Relative organ weights of placenta, fetal liver, and fetal brain were normalized to their respective fetal body weight (g) and expressed as organ weight/body weight ratios. Fetuses were obtained from animals exposed to normoxia (NMX) or hypoxia (HPX) in the presence or absence of N-acetylcysteine (NAC). Data are mean ± standard error of the mean (SEM), ***P < .001, **P < .01 (NMX n = 8, HPX n = 9, HPX + NAC n = 8, and NMX + NAC n = 10).

Figure 2.

Malondialdehyde (MDA) levels (nmoles/mg protein) of the fetal guinea pig liver, heart, lung, and brain. Fetal tissues were obtained from animals exposed to normoxia (NMX) or hypoxia (HPX) in the presence or absence of N-acetylcysteine (NAC). Data are mean ± standard error of the mean (SEM), **P < .01, *P < .05 (NMX n = 5, HPX n = 4-5, HPX + NAC n = 5-6, NMX + NAC n = 5-6).

Figure 3.

Cytochrome oxidase (CCO) activity of fetal guinea pig liver, heart, lung, and brain of animals exposed to normoxia (NMX) or hypoxia (HPX) in the presence or absence of N-acetylcysteine (NAC). Data are mean ± standard error of the mean (SEM) and expressed as CCO-specific activity (1 U = 1 μmol of oxidized cytochrome c/min per mg protein), *P < .05. Fetal liver, lung, and brain NMX n = 8-12, HPX n = 9-12, HPX + NAC n = 8-12, NMX + NAC n = 10-12. Fetal heart (n = 5 for each group).

Figure 4.

Western blot analysis of cytochrome oxidase subunit 4 (COX4) protein expression of fetal guinea pig liver, heart, lung, and brain of animals exposed to normoxia (NMX) or hypoxia (HPX) in the presence or absence of N-acetylcysteine (NAC). Data are mean ± standard error of the mean (SEM). The COX4 expression is measured as a ratio of COX4 signal/Coomassie stain. *P < .05. Molecular weight (MW) = 16 kDa. NMX n = 3-4, HPX n = 3-4, HPX + NAC n = 3-4, and NMX + NAC n = 3-4.