Extracellular Superoxide Production Research Articles

Extra-cellular superoxide promotes T cell expansion through inactivation of nitric oxide

The mechanism and regulation of immunosuppression by nitric oxide (NO) is unclear. Extra-cellular superoxide (EC-O 2 −) production by NADPH-oxidase (phox) may prevent NO-mediated suppression of T cell proliferation. p47 phox−/− mice are resistant to experimental allergic encephalomyelitis (EAE), coinciding with enhanced splenic NO activity, but no causal link was established. Here, we demonstrate such link, since p47 phox−/− mice developed severe EAE by adoptive transfer, but only if NO production during ex vivo donor cell reactivation was inhibited. EC-O 2 − production increased during cognate T cell reactivation, while inhibition of EC-O 2 − by exogenous superoxide dismutase enhanced NO activity. By inhibiting NO, EC-O 2 − production promotes T cell expansion during peripheral immune-response activation, not during tissue inflammation.

Dietary brewers yeast and the prebiotic Grobiotic™AE influence growth performance, immune responses and resistance of hybrid striped bass ( Morone chrysops× M. saxatilis) to Streptococcus iniae infection

Use of prebiotics, nondigestible dietary ingredients that beneficially affect the host by selectively stimulating the growth of and/or activating the metabolism of health-promoting bacteria in the intestinal tract, is a novel concept in aquaculture. Two separate feeding trials were conducted to evaluate graded levels of a commercial prebiotic Grobiotic™AE, a mixture of partially autolyzed brewers yeast, dairy ingredient components and dried fermentation products, in the diet of hybrid striped bass, as compared to partially autolyzed brewers yeast (Brewtech®). The basal diet in both trials was formulated to contain 40% protein, 10% lipid and an estimated digestible energy level of 3.5 kcal/g. Two levels (1% and 2% of diet) of Grobiotic™AE and brewers yeast were added to the basal diet with menhaden fish meal and menhaden oil adjusted to provide isonitrogenous and isolipidic diets. Each diet was fed to five (trial 1) or three (trial 2) replicate groups of juvenile hybrid striped bass in 110-l aquaria twice daily at rates approximating apparent satiation for 7 weeks (trial 1) or 4 weeks (trial 2). Enhanced growth performance was generally observed in fish fed the diets supplemented with Grobiotic™AE or brewers yeast compared to the basal diet after 7 weeks of feeding in trial 1. Significantly higher ( P<0.05) feed efficiency was observed in fish fed diets supplemented with 1% and 2% Grobiotic™AE. After 4 weeks of feeding in trial 2, growth and feed efficiency were not significantly affected by the various dietary treatments, although some immunological responses were altered. Neutrophil oxidative radical anion production and intracellular superoxide anion production of head kidney macrophages tended to be higher in fish fed diets supplemented with brewers yeast or Grobiotic™AE, while extracellular superoxide anion production of head kidney macrophages from fish fed diets with 1% and 2% brewers yeast and 1% Grobiotic™AE was significantly ( P<0.01) higher than that of fish fed the basal diet. All groups of fish fed brewers yeast and Grobiotic™AE showed a significantly ( P<0.01) enhanced survival (73.3–90%) after bath exposure to Streptococcus iniae compared to fish fed the basal diet (53.3%). Based on these data, it is concluded that Grobiotic™AE and a partially autolyzed brewers yeast can serve as functional feedstuffs in the diets of hybrid striped bass by enhancing growth performance and immunological responses. Further research is needed into the mechanism(s) of action for prebiotics such as Grobiotic™AE and their application in aquaculture.

High rates of extracellular superoxide production by lichens in the suborder Peltigerineae correlate with indices of high metabolic activity

ABSTRACTRates of extracellular superoxide radical (O2· −) formation were measured in 34 species of lichens from different taxonomic groupings and contrasting habitats before and after desiccation stress. All 21 species from the suborder Peltigerineae produce O2· − extracellularly at high rates, even when they are not stressed. In addition, some species show a burst of O2· − production during rehydration following desiccation. Extracellular production of O2· − is almost absent in the species from other lichen groups. In general, production of high levels of O2· − and the existence of an inducible oxidative burst are best developed in species growing in wet microhabitats. Rates of O2· − production are also positively correlated to previously published indices of lichen metabolic activity. Preliminary studies on the identity of the O2· − producing enzymes suggest that they do not possess the classical characteristics of those suggested to produce reactive oxygen species in higher plants. Patterns of O2· − production are discussed in terms of the strategies used by different lichens groups in their defence against pathogenic fungi and bacteria.

Critical role of microglial NADPH oxidase-derived free radicals in the in vitro MPTP model of Parkinson's disease.

1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) damages dopaminergic neurons as seen in Parkinson's disease. Although increasing evidence suggests an involvement of glia in MPTP neurotoxicity, the nature of this involvement remains unclear. Exploiting the advantage of cell culture systems, we demonstrated that microglia, but not astroglia, significantly enhanced the progression of MPTP-induced dopaminergic neurodegeneration. Characterization of the temporal relationship between neurodegeneration and microglial activation demonstrates that reactive microgliosis resulting from MPTP-initiated neuronal injury, but not direct activation, underlies the microglia-enhanced MPTP neurotoxicity. Mechanistically, through the release of NADPH oxidase-derived reactive oxygen species, microglia contribute to the progressive neuronal damage. Among the factors measured, the production of extracellular superoxide was the most prominent. NADPH oxidase inhibitor, apocynin, attenuated MPTP-induced dopaminergic neurodegeneration only in the presence of glia. More importantly, dopaminergic neurons from mice lacking NADPH oxidase, a key enzyme for superoxide production in immune cells, are significantly more resistant to MPTP neurotoxicity than those from wild-type controls, and microglia dictate the resistance. This study demonstrates that reactive microgliosis triggered by MPTP-induced neuronal injury and NADPH oxidase-mediated superoxide production in microglia constitute an integral component of MPTP neurotoxicity. This study also suggests that NADPH oxidase may be a promising target for therapeutic interventions in Parkinson's disease.

Evaluation of brewers yeast ( Saccharomyces cerevisiae) as a feed supplement for hybrid striped bass ( Morone chrysops× M. saxatilis)

Two separate feeding trials were conducted to evaluate graded levels of dried brewers yeast in the diet of hybrid striped bass. A basal diet was formulated to contain 40% protein, 10% lipid and an estimated digestible energy level of 3.5 kcal/g. In Trial 1, three incremental levels of dried brewers yeast (1%, 2% and 4% of diet) were added to the basal diet in place of cellulose. In Trial 2, the same levels of brewers yeast were added to the basal diet, but menhaden fish meal and menhaden oil were adjusted to provide isonitrogenous and isolipidic diets. Each diet was fed to three (Trial 1) or four (Trial 2) replicate groups of juvenile hybrid striped bass twice daily at rates approximating apparent satiation for 6 or 8 weeks. After the second feeding trial, a Streptococcus iniae bath challenge was executed to test the effects of diet on disease resistance. Enhanced weight gain and feed efficiency were generally observed in fish fed the diets supplemented with yeast compared to the basal diet in both trials. In the second trial, body composition of whole fish, hemocrit and serum lysozyme levels were observed to be within normal ranges and not influenced by the various dietary treatments. After 9 weeks of feeding in the second trial, exposure to S. iniae resulted in no mortality and reduced signs of disease in fish fed diets supplemented with 2% and 4% brewers yeast, while 20% mortality was observed in fish fed the control diet ( P=0.1). In the second trial, blood neutrophil oxidative radical production, extracellular and intracellular superoxide anion production of head kidney macrophages and serum lysozyme were measured after 16 weeks of feeding each diet. Fish fed the diet with 2% brewers yeast were found to have significantly ( P<0.01) higher blood neutrophil oxidative radical and extracellular superoxide anion production of head kidney macrophages than control fish. However, no significant differences in intracellular superoxide anion and serum lysozyme were observed among the treatments. Based on the result of this study, it is concluded that brewers yeast positively influenced growth performance and feed efficiency of hybrid striped bass as well as resistance to S. iniae infection. In addition, results of immune response assays demonstrate that brewers yeast can be administered for relatively long periods without causing immunosuppression.

Wounding induces a burst of extracellular superoxide production in Peltigera canina

Wounding induces a burst of extracellular superoxide production in Peltigera canina

Increased reactive oxygen species production with antisense oligonucleotides directed against uncoupling protein 2 in murine endothelial cells.

Uncoupling protein 2 (UCP-2) belongs to the mitochondrial anion carrier family. It is ubiquitously expressed but is most abdundant in the reticuloendothelial system. In addition to uncoupling function, UCP-2 modulates the production of reactive oxygen species (ROS) by isolated mitochondria. Using an antisense oligonucleotide strategy, we investigated whether a defect in UCP-2 expression modulates ROS in intact endothelial cells. Murine endothelial cells (CRL 2181) pretreated by antisense oligonucleotides directed against UCP-2 mRNA exhibited a significant and specific increase in membrane potential and intracellular ROS level compared with control scrambled or anti-UCP-1 and -UCP-3 antisense oligonucleotides. These specific changes induced by UCP-2 antisense oligonucleotides were correlated with a rise in extracellular superoxide anion production and oxidative stress assessed by thiobarbituric acid reactive substance values. Taken together, these data suggest a role for UCP-2 in control of ROS production and subsequent oxidation of surrounding compounds mediating oxidative stress of endothelial cells. These data also support the notion that manipulations of UCP-2 at the genetic level could control ROS metabolism at the cellular level.

Extracellular superoxide production by Enterococcus faecalis requires demethylmenaquinone and is attenuated by functional terminal quinol oxidases.

The intestinal commensal bacterium, Enterococcus faecalis, is unusual among prokaryotic organisms in its ability to produce substantial extracellular superoxide. Transposon mutagenesis, allelic replacement, and electron spin resonance (ESR)-spin trapping showed that superoxide production and generation of derivative hydroxyl radical were dependent on membrane-associated demethylmenaquinone. Extracellular superoxide was generated through univalent reduction of oxygen by reduced demethylmenaquinone. Moreover, extracellular superoxide production was inhibited by exogenous haematin, an essential cofactor for cytochrome bd, and by fumarate, a substrate for fumarate reductase. As integral membrane quinol oxidases, cytochrome bd and fumarate reductase redox cycle demethylmenaquinone, and are necessary for aerobic and anaerobic respiration respectively. A rat model of intestinal colonization demonstrated that conditions exist in the mammalian intestinal tract that permit a mode of respiration for E. faecalis that results in the formation of hydroxyl radical. These results identify and characterize the mechanism by which E. faecalis generates extracellular free radicals.

High rates of extracellular superoxide production in bryophytes and lichens, and an oxidative burst in response to rehydration following desiccation

Summary The mechanism of extracellular superoxide radical (O2·−) formation and the role of the oxidative burst in response to desiccation stress is reported here in bryophytes and lichens from habitats of contrasting water availability. Rates of extracellular production of O2·− radicals were measured, before and after desiccation stress, by the oxidation of epinephrine to adrenochrome, determined spectrophotometrically. Several desiccation‐sensitive lichens and bryophytes that grow in very wet microhabitats produced O2·− extracellularly at high rates, even when they were not stressed. In addition, some species showed a powerful burst of O2·− production during rehydration following desiccation. Production of high levels of O2·− and the existence of an inducible oxidative burst was best developed in cyanobacterial lichens, a hornwort and the two thalloid liverworts tested. Extracellular production of O2·− was almost absent from all mosses tested, a leafy liverwort, a filmy fern and from desiccation‐tolerant lichens. Patterns of O2·− production are discussed in terms of their possible role as a defence against pathogenic fungi and bacteria.

Ras oncogene expression determines sensitivity for intercellular induction of apoptosis.

Fibroblasts carrying an inducible ras oncogene acquire the transformed phenotype after oncogene induction. As a consequence, the transformed cells become sensitive to intercellular induction of apoptosis, a novel regulatory process directed by non-transformed fibroblasts against their transformed descendants. The causal relationship between oncogene expression and sensitivity to intercellular induction of apoptosis is based on extracellular superoxide anion production by oncogene-expressing cells. Superoxide anions (after dismutation to hydrogen peroxide) thereby foster HOCl synthesis and at the same time direct the selectivity of apoptosis induction through hydroxyl generation from HOCl. In parallel, ras expression enhances the sensitivity of fibroblasts for apoptosis-inducing stimuli like cycloheximide, ceramide and mitomycin C. This sensitization seems to be based on a decreased concentration of short lived endogenous apoptosis inhibitors. TGF-beta, like ras induction, decreases the concentration of endogenous apoptosis inhibitors, but does not induce the transformed phenotype. Therefore, TGF-beta treatment alone is not sufficient to render fibroblasts sensitive for intercellular induction of apoptosis, but TGF-beta treatment in parallel with ras activation enhances intercellular induction of apoptosis. Our findings demonstrate that Ras-mediated superoxide anion production determines sensitivity to intercellular induction of apoptosis, whereas the parallel decrease in endogenous apoptosis inhibitors modulates the kinetics of apoptosis induction.

Phenylethylamine-induced generation of reactive oxygen species and ascorbate free radicals in tobacco suspension culture: mechanism for oxidative burst mediating Ca2+ influx.

In the previous paper [Kawano et al. (2000a) Plant Cell Physiol. 41: 1251], we demonstrated that addition of phenylethylamine (PEA) and benzylamine can induce an immediate and transient burst of active oxygen species (AOS) in tobacco suspension culture. Detected AOS include H2O2, superoxide anion and hydroxyl radicals. Use of several inhibitors suggested the presence of monoamine oxidase-like H2O2-generating activity in the cellular soluble fraction. It was also suggested that peroxidase(s) or copper amine oxidase(s) are involved in the extracellular superoxide production as a consequence of H2O2 production. Since more than 85% of the PEA-dependent AOS generating activity was localized in the extracellular space (extracellular fluid + cell wall), extracellularly secreted enzymes, probably peroxidases, may largely contribute to the oxidative burst induced by PEA. The PEA-induced AOS generation was also observed in the horseradish peroxidase (HRP) reaction mixture, supporting the hypothesis that peroxidases catalyze the oxidation of PEA leading to AOS generation. In addition to AOS production, we observed that PEA induced an increase in monodehydroascorbate radicals (MDA) in the cell suspension culture and in HRP reaction mixture using electron spin resonance spectroscopy and the newly invented MDA reductase-coupled method. Here we report that MDA production is an indicator of peroxidase-mediated generation of PEA radical species in tobacco suspension culture.

Intra- and extracellular measurement of reactive oxygen species produced during heat stress in diaphragm muscle.

Skeletal muscles are exposed to increased temperatures during intense exercise, particularly in high environmental temperatures. We hypothesized that heat may directly stimulate the reactive oxygen species (ROS) formation in diaphragm (one kind of skeletal muscle) and thus potentially play a role in contractile and metabolic activity. Laser scan confocal microscopy was used to study the conversion of hydroethidine (a probe for intracellular ROS) to ethidium (ET) in mouse diaphragm. During a 30-min period, heat (42 degrees C) increased ET fluorescence by 24 +/- 4%, whereas in control (37 degrees C), fluorescence decreased by 8 +/- 1% compared with baseline (P < 0.001). The superoxide scavenger Tiron (10 mM) abolished the rise in intracellular fluorescence, whereas extracellular superoxide dismutase (SOD; 5,000 U/ml) had no significant effect. Reduction of oxidized cytochrome c was used to detect extracellular ROS in rat diaphragm. After 45 min, 53 +/- 7 nmol cytochrome c. g dry wt(-1). ml(-1) were reduced in heat compared with 22 +/- 13 nmol. g(-1). ml(-1) in controls (P < 0.001). SOD decreased cytochrome c reduction in heat to control levels. The results suggest that heat stress stimulates intracellular and extracellular superoxide production, which may contribute to the physiological responses to severe exercise or the pathology of heat shock.

Changes with aging in the modulation of macrophages by norepinephrine

The effect of aging on the norepinephrine (NE)-induced modulation of phagocytic and oxygen-dependent microbicidal processes of mouse peritoneal macrophages was studied. Phagocytosis of latex beads on culture plates and superoxide anion production was evaluated in young (12 weeks), adult (22 weeks), mature (48 weeks) and old (72 weeks) BALB/c mice after in vitro incubation with 10 −12, 10 −9, 10 −7, 10 −5 or 10 −3 M concentrations of NE. The results indicate that the phagocytic response to NE is quite similar in young and mature mice, with increased phagocytosis after incubation with 10 −7 and 10 −3 M, and decreased phagocytosis with 10 −5 M. Macrophages from adult mice increased their phagocytic capacity after incubation with the highest concentrations of NE (10 −5 and 10 −3 M) and macrophages from old animals only were stimulated with 10 −3 M. In addition, it was found that usually NE increased the extracellular superoxide anion production in the absence of phagocytosis in adult mice. No statistically significant changes were found in intracellular superoxide anion levels, but an increase was seen after phagocytosis in macrophages from adult, mature and old animals, especially after incubation with 10 −5 M. In conclusion, the results obtained indicate that the modulation of macrophages by NE does not only depend on the concentration of this neurotransmitter, but also on age.

Enterococcus faecalis bearing aggregation substance is resistant to killing by human neutrophils despite phagocytosis and neutrophil activation.

Enterococcus faecalis aggregation substance (AS) mediates efficient bacterium-bacterium contact to facilitate plasmid exchange as part of a bacterial sex pheromone system. We have previously determined that AS promotes direct, opsonin-independent binding of E. faecalis to human neutrophils (PMNs) via complement receptor type 3 and other receptors on the PMN surface. We have now examined the functional consequences of this bacterium-host cell interaction. AS-bearing E. faecalis was phagocytosed and internalized by PMNs, as determined by deconvolution fluorescence microscopy. However, these bacteria were not killed by PMNs, and internalized bacteria excluded propidium iodide, indicating intact bacterial membranes. Resistance to killing occurred despite activation of PMNs, as indicated by an increase in both functional and total surface Mac-1 expression, shedding of L-selectin, and an increase in PMN extracellular superoxide and phagosomal oxidant production. Deconvolution fluorescence microscopy also revealed that phagosomes containing AS-bearing bacteria were markedly larger than phagosomes containing opsonized E. faecalis, suggesting that some modification of phagosomal maturation may be involved in AS-induced resistance to killing. PMN phagosomal pH was significantly higher after ingestion of nonopsonized AS-bearing E. faecalis than after that of opsonized bacteria. The novel ability of AS to promote intracellular survival of E. faecalis inside PMNs suggests that AS may be a virulence factor used by strains of E. faecalis.

In vivo survival of Enterococcus faecalis is enhanced by extracellular superoxide production.

Enterococci are major nosocomial pathogens comprising 9% of the over 100,000 primary hospital-acquired bacteremias that occur annually in the United States (4). Despite the significant morbidity and mortality caused by these nosocomial pathogens, little is known of enterococcal traits that promote infection. Recent observations by us indicate that most Enterococcus faecalis and a few E. faecium strains generate substantial extracellular Superoxide (ESO), with invasive isolates having significantly greater production than commensal strains (6). In this study we investigated the role of ESO production in subcutaneous infection using a previously described murine model (5).KeywordsAromatic Amino AcidEnterococcus FaecalisInvasive IsolateSubcutaneous InfectionOklahoma HealthThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Enhancement of Superoxide Production and Protection against Heat Shock by HSP27 in Fibroblasts

Extracellular superoxide production in the Chinese hamster CCL39 fibroblast cell line was detected by direct counting of luminol-enhanced chemiluminescence in the presence of horseradish peroxidase (HRP). This superoxide production, which is directly related to cell density, can be inhibited by diphenylene iodonium, a specific inhibitor of NADPH-oxidase. The O−2generation is transiently inhibited following exposure of the cells to elevated temperatures. To study the possible relationship between HSP27 and its modulation of NADPH-oxidase activity we measured the generation of O−2activity in CCL39 cells containing either the gene coding for human HSP27 or a mutant form of this gene leading to HSP27 unable to be phosphorylated. Cells with the enhanced expression of normal human HSP27 as well as the non-phosphorylatable protein exhibited a 20-fold higher superoxide production than control CCL39 cells. Furthermore, we demonstrate that accumulation of normal HSP27 appears to play a role in prevention of NADPH-oxidase inhibition by heat.

Augmented production of extracellular superoxide by blood isolates of Enterococcus faecalis.

To assess the frequency of extracellular superoxide (O-2) production by enterococci, multiple species were surveyed in a whole organism assay for their ability to reduce ferricytochrome c in a superoxide dismutase-inhibitable fashion. For stool and clinical enterococcal isolates and 12 type strains, only Enterococcus faecalis (87/91 isolates and ATCC 19433), Enterococcus faecium (5/13 isolates), Enterococcus casseliflavus (ATCC 25778), and Enterococcus gallinarum (ATCC 35038) produced extracellular O-2. Among 106 strains comprising 13 species of enteric gram-negative bacilli and gram-positive cocci, only Lactococcus lactis subspecies lactis produced extracellular O-2. Mean (+/-SE) rates of extracellular O-2 production in vitro by E . faecalis for isolates associated with bacteremia and endocarditis and for isolates from stool were 2.4+/-0.2, 1.9+/-0.2, 1.5+/-0.3 nmol of O-2 min(-1) 10(9) cfu(-1), respectively (P=.025, analysis of variance), suggesting an association between invasiveness and extracellular O-2 production.

Evaluation of Oxidative Processes in Human Pigment Epithelial Cells Associated with Retinal Outer Segment Phagocytosis

To investigate the nature of the oxidative event that occurs during phagocytosis of retinal outer segments (ROS) by cultured human retinal pigment epithelial (RPE) cells, cells were incubated with isolated bovine ROS labeled with either the fluorescent probe carboxy-SNAFL-2 or the nonfluorescent, oxidizable probe 2′,7′dichlorodihydrofluorescein (H2DCF). The increase in fluorescence following phagocytosis was measured by a flow cytometer. Other measurements included: oxygen consumption using a Clark-type oxygen electrode, extracellular superoxide release by superoxide dismutase inhibitable lucigenin chemiluminescence, intracellular hydrogen peroxide (H2O2) production, and the effect of catalase inhibition on cellular thiobarbituric acid-reactive substances (TBARS) caused by phagocytosis. The activities of the enzymes NADPH oxidase and palmitoyl-CoA oxidase were also measured. H2DCF attached to bovine ROS was oxidized during phagocytosis with a time course suggesting oxidation subsequent to ROS uptake. Measurements of oxygen consumption showed a time-dependent increase of 10%, 4 h after ROS feeding, attributable to a doubling of the cyanide-resistant oxygen consumption. Intracellular H2O2 production also doubled 4 h after ROS phagocytosis. ROS uptake by RPE cells produced no significant extracellular superoxide, while extracellular superoxide production was readily demonstrated in a control macrophage cell line. Enzyme activity measurements showed that incubation of RPE cells with ROS doubled catalase activity without affecting superoxide dismutase or glutathione peroxidase activities. Inhibition of catalase during ROS uptake increased TBARS by 66%. Other enzyme activity measurements showed that human RPE cells possess both NADPH oxidase and palmitoyl-CoA oxidase activities. We conclude that ROS phagocytosis subjects RPE cells to an oxidative event on the same order of magnitude as measured in a macrophage. The event is not an extracellular macrophage-type respiratory burst and may be due to intracellular H2O2 resulting from an NADPH oxidase in the phagosome or from β-oxidation of ROS lipids in peroxisomes. Irrespective of case, the enzyme catalase appears to be essential in protecting the RPE cell against reactive oxygen species produced during phagocytosis.

OXIDATIVE STRESS MEDIATES TUMOR PROMOTER-INDUCED PROLIFERIN GENE-EXPRESSION IN C3H10T1/2 CELLS

Increased expression of the proliferin gene family occurs upon exposure of C3H10T1/2 cells to a broad range of chemical agents known to promote morphological transformation and likely to increase cellular reactive oxygen species. To determine if proliferin gene expression is influenced by reactive oxygen species, superoxide radicals were generated in culture by the xanthine/xanthine oxidase couple. The 1 kb cytoplasmic proliferin transcript accumulated up to ten-fold following extracellular superoxide production. Within certain concentration ranges of catalase and superoxide dismutase activity, xanthine oxidase-induced proliferin expression was reduced to control levels, while expression was increased at other concentrations. Induction of proliferin by the tumour promoters butylated hydroxytoluene or TPA was efficiently inhibited at certain concentrations of catalase and superoxide dismutase, but retinoic acid had no effect. Proliferin induction by a recently identified promoter of transformation, tri-n-butyltin chloride, was stimulated by catalase, superoxide dismutase and retinoic acid, but inhibited at higher concentrations of N-acetyl cysteine. c-fos preceded proliferin induction by butylated hydroxytoluene, but several other oncogenes and growth-regulated genes were unaffected. The results support a mechanism for tumour promoter-induced proliferin gene expression that involves a response to superoxide anions, hydrogen peroxide or other shifts in the cellular equilibrium between pro-oxidant and anti-oxidant moieties.

Superoxide dismutase does not prevent delayed hypoperfusion after incomplete cerebral ischaemia in the rat

Local cerebral blood flow (1CBF) was measured autoradiographically 60 minutes after 15 minutes of forebrain ischaemia in rats treated with superoxide dismutase (SOD) before (50 mg.kg-1 body weight) or at the end of the ischaemia period (4 mg.kg-1 body weight). Incomplete forebrain ischaemia was produced by a combination of common carotid artery occlusion and bleeding to a mean arterial blood pressure of 50 mmHg. During ischaemia the 1CBF values in cortical areas were less than 3% of the preischaemic values and treatment with SOD prior to ischaemia did not influence 1CBF during ischaemia. Sixty minutes after termination of cerebral ischaemia the 1CBF values were decreased to between 40 and 60% of values found in control animals. Neither form of treatment improved the postischaemic cerebral blood flows. The results imply that postischaemic flow disturbances in the brain may not be due to extracellular superoxide production.