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.

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Intrauterine hypoxia impacts fetal growth and organ function. Inducible nitric oxide synthase (iNOS) and neuronal NOS (nNOS) expression was measured to assess the response of fetal hearts to hypoxic (HPX) stress. Pregnant guinea pigs were housed in a hypoxic chamber (10.5% O 2 for 14 d, n ϭ 17) or room air [normoxic (NMX), n ϭ 17͔. Hearts of anesthetized near-term fetuses were removed. mRNA ͓hypoxia-inducible factor, (HIF)-1␣, 1␤, 2␣, 3␣, iNOS, and nNOS͔ and protein levels (HIF-1␣, iNOS, and nNOS) of fetal cardiac left ventricles were quantified by real time polymerase chain reaction (PCR) and Western analysis, respectively. Cardiac nitrite/nitrate levels were measured in the presence/absence of L-N 6 -(1-iminoethyl)-lysine (L-NIL), an iNOS inhibitor, administered to pregnant sows. Hypoxia significantly increased fetal cardiac HIF-1␣ and -2␣ mRNA, HIF-1␣ protein but not HIF-3␣ or -1␤ mRNA levels. Hypoxia increased both iNOS mRNA (by 5ϫ) and protein (by 23%) levels but had no effect on nNOS levels. Nitrite/nitrate levels were increased in HPX hearts by 2.5ϫ and decreased with L-NIL by 67 Ϯ 14%. Thus, up-regulation of iNOS-derived nitric oxide (NO) generation is an important mechanism by which fetal hearts respond to chronic hypoxic stress. T he adaptive response of the fetal heart to intrauterine stress is critical for its survival. Several studies using high altitude (1,2) exposure to acute and chronic hypoxia (2,3), and anemia (4) have demonstrated how the fetal cardiovascular system responds to hypoxic (HPX) stress. Depending on the severity and duration of the HPX conditions, as well as the gestational age of the fetus, cardiac adaptations have been associated with altered coronary blood flow (2,5,6), increased heart size (1), and decreased contractile performance (7). The underlying mechanisms mediating these changes in fetal heart morphology and function are not fully understood.Hypoxia is a potent stimulus for gene activation of several genes (8), including nitric oxide synthase (NOS), the synthetic enzyme that generates nitric oxide (NO) from L-arginine oxidation (9). NO is derived from three isoforms of NOS ͓endothelial NOS (eNOS); neuronal NOS (nNOS); and inducible NOS (iNOS)͔, all of which are expressed in the heart (10,11). Specifically, eNOS is expressed constitutively in both endothelial cells and cardiomyocytes (10), nNOS in both cardiomyocytes and conducting tissue of cardiac ventricles (12), and iNOS in cardiomyocytes (12) and in mature hearts under conditions of hypoxia (12-14), heart failure (15), left ventricular hypertrophy (16) and cardiac cyanosis in children (17). NOS gene expression has been reported to be oxygensensitive, with levels varying among cardiac cells (18,19) and endothelial cells of differing vascular origin (20 -22). For example, hypoxia increases eNOS expression in porcine coronary artery endothelial cells (20), newborn (23-25) and adult rabbit heart ventricles (26), and adult guinea pig ventricles (27), and decreases expression in pulmonary artery endothelial cell...

Chronic exposure to hypoxia during pregnancy generates a stressed intrauterine environment that may lead to fetal organ damage. The objectives of the study are (1) to quantify the effect of chronic hypoxia in the generation of oxidative stress in fetal guinea pig liver and (2) to test the protective effect of antioxidant treatment in hypoxic fetal liver injury. Pregnant guinea pigs were exposed to either normoxia (NMX) or 10.5% O 2 (HPX, 14 days) prior to term (65 days) and orally administered N-acetylcysteine ([NAC] 10 days). Near-term anesthetized fetuses were excised and livers examined by histology and assayed for malondialdehyde (MDA) and DNA fragmentation. Chronic HPX increased erythroid precursors, MDA (NMX vs HPX; 1.26 + 0.07 vs 1.78 + 0.07 nmol/mg protein; P < .001, mean + standard error of the mean [SEM]) and DNA fragmentation levels in fetal livers (0.069 + 0.01 vs 0.11 + 0.005 OD/mg protein; P < .01). N-acetylcysteine inhibited erythroid aggregation and reduced (P < .05) both MDA and DNA fragmentation of fetal HPX livers. Thus, chronic intrauterine hypoxia generates cell and nuclear damage in the fetal guinea pig liver. Maternal NAC inhibited the adverse effects of fetal liver damage suggestive of oxidative stress. The suppressive effect of maternal NAC may implicate the protective role of antioxidants in the prevention of liver injury in the hypoxic fetus.

This study tested the hypothesis that maternal nicotine ingestion increases matrix metalloproteinase (MMP) expression in fetal hearts, which is mediated by the generation of reactive oxygen species. Timed pregnant guinea pigs were administered either water alone, nicotine (200 mg/mL), N-acetylcysteine (NAC), or nicotine plus NAC in their drinking water for 10 days at 52-day gestation (term ¼ 65 days). Near-term (62 days), anesthetized fetuses were extracted, hearts were excised, and left cardiac ventricles snap frozen for analysis of MMP-2/-9/-13 protein and activity levels. Interstitial collagens were identified by Picrosirius red stain to assess changes in the extracellular matrix. Prenatal nicotine increased active MMP-2 forms and interstitial collagen but had no effect on either pro-or active MMP-9 or MMP-13 forms. In the presence of nicotine, NAC decreased active MMP-2 protein levels and reversed the nicotine-induced increase in collagen staining. We conclude that prenatal nicotine alters MMP-2 expression in fetal hearts that may be mediated by reactive oxygen species generation.