Manipulation of Antioxidant Pathways in Neonatal Murine Brain

Abstract

To assess the role of brain antioxidant capacity in the pathogenesis of neonatal hypoxic-ischemic brain injury, we measured the activity of glutathione peroxidase (GPX) in both humansuperoxide dismutase-1 (hSOD1) and human-GPX1 overexpressing transgenic (Tg) mice after neonatal hypoxia-ischemia (HI). We have previously shown that mice that overexpress the hSOD1 gene are more injured than their wild-type (WT) littermates after HI, and that H 2 O 2 accumulates in HI hSOD1-Tg hippocampus. We hypothesized that lower GPX activity is responsible for the accumulation of H 2 O 2 . Therefore, increasing the activity of this enzyme through gene manipulation should be protective. We show that brains of hGPX1-Tg mice, in contrast to those of hSOD-Tg, have less injury after HI than WT littermates: hGPX1-Tg, median injury score ϭ 8 (range, 0 -24) versus WT, median injury score ϭ 17 (range, 2-24), p Ͻ 0.01. GPX activity in hSOD1-Tg mice, 2 h and 24 h after HI, showed a delayed and bilateral decline in the cortex 24 h after HI (36.0 Ϯ 1.2 U/mg in naive hSOD1-Tg versus 29.1 Ϯ 1.7 U/mg in HI cortex and 29.2 Ϯ 2.0 for hypoxic cortex, p Ͻ 0.006). On the other hand, GPX activity in hGPX1-Tg after HI showed a significant increase by 24 h in the cortex ipsilateral to the injury (48.5 Ϯ 5.2 U/mg, compared with 37.2 Ϯ 1.5 U/mg in naive hGPX1-Tg cortex, p Ͻ 0.008). These findings support the hypothesis that the immature brain has limited GPX activity and is more susceptible to oxidative damage and may explain the paradoxical effect seen in ischemic neonatal brain when SOD1 is overexpressed. (Pediatr Res 56: 656-662, 2004) Abbreviations CCA, common carotid artery GPX, glutathione peroxidase GSH, glutathione HI, hypoxia-ischemia P, postnatal day red-ox, oxidation-reduction reaction ROS, reactive oxygen species SOD1, copper-zinc superoxide dismutase Tg, transgenic WT, wild-typeThe developing brain faces unique challenges from oxidative stress, and has responses to injury and protective mechanisms that are different from the mature nervous system. The developmental profile of the activity of the endogenous antioxidant enzymes SOD1 and selenium-dependent GPX have been described for both rats (1-3) and mice (4). In the mouse brain, GPX activity is almost exclusively due to the classic, cellular glutathione peroxidase GPX1 (EC 1.11.1.9) (5). In the CD1 mouse, the outbred background strain used in this study, SOD1 activity decreases around the time of birth, and then remains unchanged to weaning. GPX activity, on the other hand, rises steeply around birth but declines sharply in the first few days of life and then remains low through weaning. Protein levels for both enzymes, however, show a steady increase during early development (4). We have shown that, in the CD1 mouse brain, adult levels of GPX activity are reached by weaning (6). Therefore, maturational differences may impact the response of the newborn brain to injury. It has been shown that the adult and juvenile brain attempts to compensate during oxidative stress in response to stroke (...

Mechanisms of Perinatal Arterial Ischemic Stroke

“…86 Several other distinct characteristics of the immature brain, such as high oxygen consumption, higher iron levels, and lower expression of several endogenous antioxidant enzymes, contribute to the higher vulnerability of the immature brain due to the deleterious actions of free radicals. 87 However, overexpression of antioxidant enzymes does not necessarily protect the neonatal brain against H-I, as was shown for superoxide dismutase overexpressing pups, and is dependent on the balance of free radical scavenging pathways 88,89 and cell types. 90 Inflammation, the third major contributor to neonatal brain injury can induce neuronal death by itself as well as by enhancing excitotoxicity and oxidative stress through the release of cytokines, free radicals, and other toxic products or trigger release of excitotoxic molecules, including glutamate.…”

Imaging selective vulnerability in the developing nervous system

“…There is an imbalance of antioxidant enzymes at this stage so that excessive H 2 O 2 is produced but overexpression of the enzyme glutathione peroxidase can compensate for this excessive accumulation (Sheldon et al 2004;LaFemina et al 2006). Low molecular weight iron is also high at this period, allowing for the generation of toxic hydroxyl radicals, but this toxicity can be partially ameliorated with desferoxamine, an iron-chelating agent (Sarco et al 2000).…”

Traumatic injury to the immature brain: Inflammation, oxidative injury, and iron-mediated damage as potential therapeutic targets

“…Such findings may be due to the regulation of antioxidant enzymes during brain development and maturation. 71 For example, metallothioneins, compounds that act as both free radical scavengers and metal chelators, are expressed at lower levels in the immature brain than in the adult and their level of expression inversely correlates with neuronal degeneration after cortical ablation injury. 72 Similarly, certain SOD isoforms are known to be expressed at lower levels in the immature brain.…”

“…78 This inability of the immature brain to respond to injury by upregulating GPx may therefore play a role in the vulnerability of the immature brain to injury. In addition, overexpression of GPx in a model of hypoxia/ischemia improves outcome, 71 suggesting that this antioxidant enzyme has great promise as a potential therapeutic agent for pediatric TBI.…”