Glutathione peroxidase, a selenium-containing enzyme, is believed to protect cells from the toxicity of hydroperoxides. The physiological role of this enzyme has previously been implicated mainly using animals fed with a selenium-deficient diet. Although selenium deficiency also affects the activity of several other cellular selenium-containing enzymes, a dramatic decrease of glutathione peroxidase activity has been postulated to play a role in the pathogenesis of a number of diseases, particularly those whose progression is associated with an overproduction of reactive oxygen species, found in selenium-deficient animals. To further clarify the physiological relevance of this enzyme, a model of mice deficient in cellular glutathione peroxidase (GSHPx-1), the major isoform of glutathione peroxidase ubiquitously expressed in all types of cells, was generated by gene-targeting technology. Mice deficient in this enzyme were apparently healthy and fertile and showed no increased sensitivity to hyperoxia. Their tissues exhibited neither a retarded rate in consuming extracellular hydrogen peroxide nor an increased content of protein carbonyl groups and lipid peroxidation compared with those of wild-type mice. However, platelets from GSHPx-1-deficient mice incubated with arachidonic acid generated less 12-hydroxyeicosatetraenoic acid and more polar products relative to control platelets at a higher concentration of arachidonic acid, presumably reflecting a decreased ability to reduce the 12-hydroperoxyeicosatetraenoic acid intermediate. These results suggest that the contribution of GSHPx-1 to the cellular antioxidant mechanism under normal animal development and physiological conditions and to the pulmonary defense against hyperoxic insult is very limited. Nevertheless, the potential antioxidant role of this enzyme in protecting cells and animals against the pathogenic effect of reactive oxygen species in other disorders remains to be defined. The knockout mouse model described in this report will also provide a new tool for future study to distinguish the physiological role of this enzyme from other selenium-containing proteins in mammals under normal and disease states.

The transcription factor NF-B, a central regulator of immunity, is subject to regulation by redox changes. We now report that cysteine-179 of the inhibitory B kinase (IKK) ␤-subunit of the IKK signalosome is a central target for oxidative inactivation by means of S-glutathionylation. S-glutathionylation of IKK-␤ Cys-179 is reversed by glutaredoxin (GRX), which restores kinase activity.

To test whether the antiapoptotic protein Bcl-2 prevents apoptosis and injury of cardiomyocytes after ischemia-reperfusion (I/R), we generated a line of transgenic mice that carried a human Bcl-2 transgene under the control of a mouse alpha-myosin heavy chain promoter. High levels of human Bcl-2 transcripts and 26-kDa Bcl-2 protein were expressed in the hearts of transgenic mice. Functional recovery of the transgenic hearts significantly improved when they were perfused as Langendorff preparations. This protection was accompanied by a threefold decrease in lactate dehydrogenase (LDH) released from the transgenic hearts. The transgenic mice were subjected to 50 min of ligation of the left descending anterior coronary artery followed by reperfusion. The infarct sizes, expressed as a percentage of the area at risk, were significantly smaller in the transgenic mice than in the nontransgenic mice (36.6 +/- 5 vs 69.9 +/- 7.3%, respectively). In hearts subjected to 30 min of coronary artery occlusion followed by 3 h of reperfusion, Bcl-2 transgenic hearts had significantly fewer terminal deoxynucleodidyl-transferase nick-end labeling-positive or in situ oligo ligation-positive myocytes and a less prominent DNA fragmentation pattern. Our results demonstrate that overexpression of Bcl-2 renders the heart more resistant to apoptosis and I/R injury.

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Summary The regulation of actin dynamics is pivotal for cellular processes such as cell adhesion, migration, and phagocytosis, and thus is crucial for neutrophils to fulfill their roles in innate immunity. Many factors have been implicated in signal-induced actin polymerization, however the essential nature of the potential negative modulators are still poorly understood. Here we report that NADPH oxidase-dependent physiologically generated reactive oxygen species (ROS) negatively regulate actin polymerization in stimulated neutrophils via driving reversible actin glutathionylation. Disruption of glutaredoxin 1 (Grx1), an enzyme that catalyzes actin deglutathionylation, increased actin glutathionylation, attenuated actin polymerization, and consequently impaired neutrophil polarization, chemotaxis, adhesion, and phagocytosis. Consistently, Grx1-deficient murine neutrophils showed impaired in vivo recruitment to sites of inflammation and reduced bactericidal capability. Together, these results present a physiological role for glutaredoxin and ROS- induced reversible actin glutathionylation in regulation of actin dynamics in neutrophils.

ABSTRACTcochlea injury, and cochlear degeneration. Such impairment produces characteristics expected of some mutations associated with age-related hearing loss and Reactive oxygen species (ROS) and oxidative stress have been implicated in cochlear injury following loud offers one possible mechanism for their action.

No abstract

Although reactive 02 species appear to participate in central nervous system (CNS) 02 toxicity, the exact roles of different reactive 02 species are undetermined. To study the contribution of extracellular superoxide anion (02 ) to CNS 02 toxicity we constructed transgenic mice overexpressing human extracellular superoxide dismutase (ECSOD; superoxide:superoxide oxidoreductase, EC 1.15.1.1) in. the brain. Remarkably, when exposed to 6 atm (1 atm = 101.3 kPA) of hyperbaric oxygen for 25 min, transgenic mice demonstrated higher mortality (83%) than nontransgenic littermates (33%; P < 0.017). Pretreatment with diethyldithiocarbamate, which inhibits both ECSOD and Cu/Zn superoxide dismutase (Cu/Zn SOD) activity, increased resistance to CNS 02 toxicity, in terms of both survival (100% in triasgenics and 93% in nontransgenics) and resistance to seizures (4-fold increase in seizure latency in both transgenic and nontransgenic mice; P < 0.05). Thus, -apparendly protects against CNS 02 toxicity. We hypothesized that°2 decreased toxicity by inactivating nitric oxide (NO'). To test this, we inhibited NO' synthase (EC 1.14.23) with N"-nitro-L-arginine to determine whether NO contributes to enhanced CNS 02 toxicity in transgenic mice. N"-nitro-L-arginine protected both transgenic and nontransgenic mice against CNS 02 toxicity (100% survival and a 4-fold delay in time to first seizure; P < 0.05), as well as abolishing the difference in sensitivity to CNS 02 toxicity between transgenic and nontransgenic mice. These results implicate NO' as an important mediator in CNS 02 toxicity and suggest that ECSOD increases CNS 02 toxicity by inhibiting 02-mediated inactivation of NO'.