Two sources of endogenous hydrogen peroxide in Escherichia coli

Mechanisms of hydrogen peroxide generation in Escherichia coli were investigated using a strain lacking scavenging enzymes. Surprisingly, the deletion of many abundant flavoenzymes that are known to autoxidize in vitro did not substantially lessen overall H₂O₂ formation. However, H₂O₂ production diminished by 25–30% when NadB turnover was eliminated. The flavin‐dependent desaturating dehydrogenase, NadB uses fumarate as an electron acceptor in anaerobic cells. Experiments showed that aerobic NadB turnover depends upon its oxidation by molecular oxygen, with H₂O₂ as a product. This reaction appears to be mechanistically adventitious. In contrast, most desaturating dehydrogenases are associated with the respiratory chain and deliver electrons to fumarate anaerobically or oxygen aerobically without the formation of toxic by‐products. Presumably, NadB can persist as an H₂O₂‐generating enzyme because its flux is limited. The anaerobic respiratory enzyme fumarate reductase uses a flavoprotein subunit that is homologous to NadB and accordingly forms substantial H₂O₂ upon aeration. This tendency is substantially suppressed by cytochrome oxidase. Thus cytochrome d oxidase, which is prevalent among anaerobes, may diminish intracellular H₂O₂ formation by the anaerobic respiratory chain, whenever these organisms encounter oxygen. These two examples reveal biochemical and physiological arrangements through which evolution has minimized the rate of intracellular oxidant formation.     

Salicylic acid,hydrogen peroxide and calcium-induced saline tolerance associated with endogenous hydrogen peroxide homeostasis in naked oat seedlings

This study investigates the role of salicylic acid (SA), hydrogen peroxide (H₂O₂) and calcium chloride (CaCl₂) singly or in combination, in inducing naked oat plant tolerance to sodium chloride (NaCl). Two-week-old naked oat plants were pretreated with both single and double of 0.5 mM SA, 0.5 mM H₂O₂ and 5 mM CaCl₂ by adding them to the culture solution for 24 h. At the end of the pretreatment, the plants were subjected to 200 mM NaCl exposure for 7 days. Data were collected on plant biomass, H₂O₂ level, antioxidant enzyme activity, non-enzymatic antioxidant content and malondialdehyde (MDA) content. Results showed that exposure to salt significantly inhibited plant growth, and the shoot and root dry weights were reduced 47.5% and 63.4%, and the H₂O₂ levels elevated 5.8 and 2.4 times in comparison with those in the control, respectively. Under the saline stress, the activities of superoxide dismutase (SOD) and catalase (CAT) were induced, but the contents of ascorbic acid (AA) and glutathione (GSH) decreased, and MDA largely accumulated. The various pretreatments efficiently counteracted the salt-caused growth inhibition, especially with H₂O₂ + CaCl₂ the shoot and root dry weights reduced only 9.4% and 24.4% of the non salt-stressed plants. The determination of endogenous H₂O₂ level demonstrated that the pretreatments induced H₂O₂ accumulation, with H₂O₂ + CaCl₂ being most efficient, but the effect was transient. After 7 days of saline stress, the H₂O₂ contents in the pretreated shoots and roots accounted for 23.7–41.8% and 31.7–57.3% of the non-pretreated plants, varying according to the different pretreatments. Under saline stress, SOD and CAT further increased, AA and GSH maintained higher levels and MDA decreased in the pretreated plants compared to the untreated plants. With application of diphenylene iodonium (DPI) during the pretreatment, which inhibited the accumulation of H₂O₂, the ameliorative effect of the pretreatment on salt-caused plant growth inhibition was reduced. However, applied DPI at the immediate end of the pretreatment did not alter its favorable role, indicating a H₂O₂ peak formed at the early time of saline stress might play an important role in regulating plant tolerance to saline stress.     

损伤条件下油菜黄单胞菜豆致病变种内源过氧化氢与间体形成的联系

【目的】本实验通过透射电子显微镜观察黄单胞菌在细胞损伤条件下的亚细胞结构和过氧化氢积累定位的变化。【方法】采用氯化铈对过氧化氢特异染色的组织化学法。【结果】细菌细胞受损伤后,出现了一个细胞壁之外的过氧化氢大量积累的额外位点。并且这个额外位点出现的频率和过氧化氢积累量都与细胞损伤的程度密切相关。另一方面,亚细胞结构的常规染色结果也显示,受到损伤的细胞中也出现一个额外的亚显微结构,即间体。间体出现的频率和大小也随着细胞损伤程度的增加而显著上升。【结论】多元线性回归分析的结果证明细胞损伤条件下细菌中出现的额外过氧化氢大量积累的位点就是间体。细胞损伤后间体中的过氧化氢积累对受损细胞应是一种主动调控机制。     

Localization changes of endogenous hydrogen peroxide during cell division cycle of Xanthomonas

Production and localization of endogenous hydrogen peroxide (H₂O₂) were investigated in strains of Xanthomonas by histochemical analysis under electron microscopy. Even though the levels of endogenous H₂O₂ production were different among various strains, the produced H₂O₂ was localized in the cell wall of all Xanthomonas strains tested. The impairment of the level of endogenous H₂O₂ accumulation resulted in a significantly decreased growth rate of bacteria, regardless if the difference of the H₂O₂ level is originally present between wild type strains or caused by mutation of the ahpC gene of Xanthomonas. The endogenous accumulation of H₂O₂ positively correlates with the cell division. Interestingly, the accumulated H₂O₂ was also localized in the mesosome-like structure and nucleoids during the cell division cycle. Furthermore, results revealed quantitative and dimensional changes of H₂O₂ accumulation in the two additional locations. These findings indicated that the additional locations of the accumulated H₂O₂ were closely associated with the process of cell division. Together, these results suggest that the endogenous H₂O₂ production plays an important role in cell proliferation of Xanthomonas.     

Accelerated hepatic haem catabolism in the selenium-deficient rat.

1. Hepatic microsomal cytochrome P-450 concentrations are lower in selenium-deficient rats treated with phenobarbital for 4 days than in similarly treated control rats. 2. No defect in haem synthesis was found on the basis of measurements of delta-aminolaevulinate synthase (EC 2.3.1.37), delta-aminolaevulinate dehydratase (EC 4.2.1.24) and ferrochelatase (EC 4.99.1.1) activities, and urinary excretion of delta-aminolaevulinate, porphobilinogen, uroporphyrin and coproporphyrin. 3. No defect in apo-(cytochrome P-450) separated by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. 4. An increase in haem catabolism was found. An 8-fold increase in hepatic microsomal haem oxygenase (EC 1.14.99.3) activity occurred in selenium-deficient rats after phenobarbital treatment, compared with a less than 2-fold increase in control rats. Also excretion of 14CO in the breath after administration of delta-amino[5-14C]laevulinate was greater by phenobarbital-treated selenium-deficient rats than by similarly treated controls. 5. These studies demonstrate that the defective induction of cytochrome P-450 by phenobarbital in selenium-deficient rats is accompanied by increased haem catabolism. This could be due to increased breakdown of cytochrome P-450 or to catabolism of haem before it attaches to the apo-cytochrome. The role of selenium in stabilizing cytochrome P-450 and/or in protecting haem from breakdown remains to be determined.     

Hematologic data of selenium-deficient and selenium-supplemented rats

Effect of dietary selenium as sodium selenite on in vivo hematological parameters of Sprague-Dawley rats was examined over a 7-month period. Dietary selenium did not alter total hemoglobin, hematocrits, erythrocyte counts, or the osmotic fragility pattern of rat blood. Selenium-excessive (1.0 ppm) rats showed slightly lower but not significantly lower methemoglobin levels than selenium-adequate (0.1 ppm) or selenium-deficient rats. Platelet counts tended to be higher in selenium-excessive rats and lower in selenium-deficient rats than in selenium-adequate rats, but the differences were not statistically significant. No clear trends were observed regarding the effect of dietary selenium on total leukocyte and differential leukocyte counts. After 7 months of dietary treatment blood glutathione peroxidase activity in selenium-deficient rats and in selenium-excessive rats was 16.8% and 142.2% of the activity in selenium-adequate rats. The results indicate that long-term selenium deficiency in rats produces no abnormal hematological parameters or any compensated hemolytic anemia in vivo.     

Cardioprotective role of endogenous hydrogen peroxide during ischemia-reperfusion injury in canine coronary microcirculation in vivo

We have recently demonstrated that endogenous H2O2 plays an important role in coronary autoregulation in vivo. However, the role of H2O2 during coronary ischemia-reperfusion (I/R) injury remains to be examined. In this study, we examined whether endogenous H2O2 also plays a protective role in coronary I/R injury in dogs in vivo. Canine subepicardial small coronary arteries (>or=100 microm) and arterioles (<100 microm) were continuously observed by an intravital microscope during coronary I/R (90/60 min) under cyclooxygenase blockade (n=50). Coronary vascular responses to endothelium-dependent vasodilators (ACh) were examined before and after I/R under the following seven conditions: control, nitric oxide (NO) synthase (NOS) inhibitor NG-monomethyl-L-arginine (L-NMMA), catalase (a decomposer of H2O2), 8-sulfophenyltheophylline (8-SPT, an adenosine receptor blocker), L-NMMA+catalase, L-NMMA+tetraethylammonium (TEA, an inhibitor of large-conductance Ca2+-sensitive potassium channels), and L-NMMA+catalase+8-SPT. Coronary I/R significantly impaired the coronary vasodilatation to ACh in both sized arteries (both P<0.01); L-NMMA reduced the small arterial vasodilatation (both P<0.01), whereas it increased (P<0.05) the ACh-induced coronary arteriolar vasodilatation associated with fluorescent H2O2 production after I/R. Catalase increased the small arterial vasodilatation (P<0.01) associated with fluorescent NO production and increased endothelial NOS expression, whereas it decreased the arteriolar response after I/R (P<0.01). L-NMMA+catalase, L-NMMA+TEA, or L-NMMA+catalase+8-SPT further decreased the coronary vasodilatation in both sized arteries (both, P<0.01). L-NMMA+catalase, L-NMMA+TEA, and L-NMMA+catalase+8-SPT significantly increased myocardial infarct area compared with the other four groups (control, L-NMMA, catalase, and 8-SPT; all, P<0.01). These results indicate that endogenous H2O2, in cooperation with NO, plays an important cardioprotective role in coronary I/R injury in vivo.     

Generation of hydrogen peroxide on oxidation of NADH by hepatic plasma membranes

The oxidation of NADH by mouse liver plasma membranes was shown to be accompanied by the formation of H₂O₂. The rate of H₂O₂ formation was less than one-tenth the rate of oxygen uptake and much slower than the rate of reduction of artificial electron acceptors. The optimum pH for this reaction was 7.0 and theK _m value for NADH was found to be 3×10^–6 M. The H₂O₂-generating system of plasma membranes was inhibited by quinacrine and azide, thus distinguishing it from similar activities in endoplasmic reticulum and mitochondria. Both NADH and NADPH served as substrates for plasma membrane H₂O₂ generation. Superoxide dismutase and adriamycin inhibited the reaction. Vanadate, known to stimulate the oxidation of NADH by plasma membranes, did not increase the formation of H₂O₂. In view of the growing evidence that H₂O₂ can be involved in metabolic control, the formation of H₂O₂ by a plasma membrane NAD(P)H oxidase system may be pertinent to control sites at the plasma membrane.     

Administration of r-VEGF-A prevents hepatic artery ligation-induced bile duct damage in bile duct ligated rats

The hepatic artery, through the peribiliary plexus, nourishes the intrahepatic biliary tree. During obstructive cholestasis, the nutritional demands of intrahepatic bile ducts are increased as a consequence of enhanced proliferation; in fact, the peribiliary plexus (PBP) displays adaptive expansion. The effects of hepatic artery ligation (HAL) on cholangiocyte functions during cholestasis are unknown, although ischemic lesions of the biliary tree complicate the course of transplanted livers and are encountered in cholangiopathies. We evaluated the effects of HAL on cholangiocyte functions in experimental cholestasis induced by bile duct ligation (BDL). By using BDL and BDL + HAL rats or BDL + HAL rats treated with recombinant-vascular endothelial growth factor-A (r-VEGF-A) for 1 wk, we evaluated liver morphology, the degree of portal inflammation and periductular fibrosis, microcirculation, cholangiocyte apoptosis, proliferation, and secretion. Microcirculation was evaluated using a scanning electron microscopy vascular corrosion cast technique. HAL induced in BDL rats 1) the disappearance of the PBP, 2) increased apoptosis and impaired cholangiocyte proliferation and secretin-stimulated ductal secretion, and 3) decreased cholangiocyte VEGF secretion. The effects of HAL on the PBP and cholangiocyte functions were prevented by r-VEGF-A, which, by maintaining the integrity of the PBP and cholangiocyte proliferation, prevents damage of bile ducts following ischemic injury.     

Selenoprotein gene expression during selenium-repletion of selenium-deficient rats

Selenium repletion of selenium-deficient rats with 20 μg selenium/kg body weight as Na₂SeO₃ was used as a model to investigate the mechanisms that control the distribution of the trace element to specific selenoproteins in liver and thyroid. Cytosolic glutathione peroxidase (cGSHPx), phospholipid hydroperoxide glutathione peroxidase (PHGSHPx), and iodothyronine 5′-deiodinase (IDI) activities were all transiently increased in liver 16 to 32 h after ip injection with selenium. However, only cGSHPx and PHGSHPx activities increased in the thyroid where IDI activity was already increased by selenium deficiency. These responses were owing to synthesis of the seleoproteins on newly synthesised and/or existing mRNAs. The selenoprotein mRNAs in the thyroid gland were increased two- and threefold after the transitory increases in selenoprotein activity. In contrast, there were parallel changes in selenoprotein mRNAs and enzyme activities in the liver, with no prolonged rises in mRNA levels. The organ differences suggest that increased thryotrophin (TSH) concentrations, which are known to induce thyrodial IDI and mRNA, may control the mRNAs for all the thyroidal selenoproteins investigated and be a major mechanism for the preservation of thyroidal selenoproteins when selenium supplies are limited.     

Stimulation of pyruvate dehydrogenase activity in adipocytes by oxytocin: Evidence for mediation of the insulin-like effect by endogenous hydrogen peroxide independent of glucose transport

Oxytocin, a nonapeptide posterior pituitary hormone, which is known to increase glucose oxidation in fat cells like insulin, is shown here to stimulate pyruvate dehydrogenase activity in these cells. The process appears to involve the activation of preexisting molecules since there was no change in the total enzyme content after full activation. The effect of oxytocin, as well as of insulin, appears to be mediated by endogenous H₂O₂ formation, as evident from (i) the enhanced [¹⁴C]formate oxidation and its greater inhibition by 3-amino-1,2,4-triazole in the hormone-treated cells than in the control. This is a measure of the catalase:H₂O₂ complex, and the dose dependence of this response is found to be identical with that of glucose oxidation via the hexose monophosphate shunt pathway and of pyruvate dehydrogenase activity; and (ii) treatment of the cells with low concentration of exogenous H₂O₂ causes the activation of pyruvate dehydrogenase to the extent which is comparable with the effect of the hormones. The ED₅₀ of oxytocin was 7 × 10^?9m, whereas the ED₅₀ of insulin was 5 × 10^?11m. The reduced, inactive (SH) derivatives of the hormones had the same dose-response relationship, but considerably lower effect (10 to 20% of the native molecules of the hormones), indicating the significant role of the disulfide bridge(s) in eliciting these metabolic responses. The stimulation of PDH by oxytocin or insulin is found to be essentially independent of medium glucose which, however, can sustain the response apparently by recycling the intracellular oxidation-reduction state. However, unlike insulin, oxytocin fails to stimulate the rapid uptake of 3-O-[³H]methyl-d-glucose in these cells. The data illustrate that the major metabolic actions of insulin, viz., glucose utilization and lipogenesis, are shared by another heterologous polypeptide hormone, e.g., oxytocin, through a common effector, H₂O₂. It is suggested that (i) oxytocin may play a limited surrogate role for insulin in these cells; and (ii) H₂O₂ production may be the general basis of oxytocin's action.     

Cyanide enhances hydrogen peroxide toxicity by recruiting endogenous iron to trigger catastrophic chromosomal fragmentation

Hydrogen peroxide (HP) or cyanide (CN) are bacteriostatic at low‐millimolar concentrations for growing Escherichia coli, whereas CN + HP mixture is strongly bactericidal. We show that this synergistic toxicity is associated with catastrophic chromosomal fragmentation. Since CN alone does not kill at any concentration, while HP alone kills at 20 mM, CN must potentiate HP poisoning. The CN + HP killing is blocked by iron chelators, suggesting Fenton's reaction. Indeed, we show that CN enhances plasmid DNA relaxation due to Fenton's reaction in vitro. However, mutants with elevated iron or HP pools are not acutely sensitive to HP‐alone treatment, suggesting that, in addition, in vivo CN recruits iron from intracellular depots. We found that part of the CN‐recruited iron pool is managed by ferritin and Dps: ferritin releases iron on cue from CN, while Dps sequesters it, quelling Fenton's reaction. We propose that disrupting intracellular iron trafficking is a common strategy employed by the immune system to kill microbes.     

Evaluation of oxidative damage in the liver of selenium-deficient rats

Previous studies have implied a relationship between Se-deficiency and oxidative stress. In the present study, the occurrence of oxidative stress due to Se-deficiency was investigated by evaluating the age dependence of growth and indices of oxidative damage for the liver of Se-deficient (SeD) rats. The ratios of liver weight to body weight of the SeD rats were greater than those of the normal rats. The values of AST and ALT (clinical indices of liver damage) were higher in the SeD rats than the normal ones especially in the young (6-12 weeks of age). The TBARS level of the 4-week-old SeD group were higher than the normal group while the level decreased with age. Conversely, the TBARS level of the normal group gradually increased and became higher than SeD group in older rats (12-20 weeks of age). Vitamin E rather than vitamin C may be consumed during oxidative stress due to Se-deficiency. Damage induced by Se-deficiency may be related to growth and the mechanisms of this damage may alter with age.     

Use of azide and hydrogen peroxide as an inhibitor for endogenous peroxidase in the immunoperoxidase method

We describe a simple and effective method for inhibition of endogenous peroxidase activity in the immunoperoxidase technique. Specimens are pre-treated with a mixture of azide and hydrogen peroxide, which is then followed by an indirect immunoperoxidase procedure. Comparison studies showed no significant loss of antigenicity or morphological details by this pre-treatment. The method is most useful for evaluating cell-specific antigens on specimens that have abundant endogenous peroxidase activity, such as blood and bone marrow.     

Proton leak and hydrogen peroxide production in liver mitochondria from energy-restricted rats

Energy restriction (ER), without malnutrition, is the only environmental intervention that consistently increases maximum life span in laboratory rodents. One theory proposes that a reduction in energy expenditure and reactive oxygen species production is the mechanism responsible for this action of ER. To further test this theory, proton leak, H2O2 production, lipid peroxidation, and protein carbonyls were measured in mitochondria from FBNF1 rats fed either a control or 40% ER diet (onset at 6 mo of age). Liver mitochondria were isolated at 7 and 12 mo of age. Liver weight decreased 25 and 36% at 1 and 6 mo of ER, respectively (P < 0.05). ER resulted in an increase (P < 0.05) in percent total polyunsaturates, n-6 polyunsaturates, and total unsaturates (6 mo only) in mitochondrial lipids. These changes, however, were not associated with significant alterations in mitochondrial function. State 4 respiration and membrane potential were not different (P > 0.05) between groups at either assessment period. Similarly, proton leak kinetics were not different between control and ER animals. Top-down metabolic control analysis and its extension, elasticity analysis, were used at the 6-mo assessment and revealed no difference in control of the oxidative phosphorylation system between control and ER rats. H2O2 production with either succinate or pyruvate/malate substrates was also not different (P > 0.05) between groups at either time point. In conclusion, ER did not alter proton leak or H2O2 production at this age or stage of restriction in liver.     

Destructive effect of anticancer oxali-palladium on heme degradation through the generation of endogenous hydrogen peroxide

The interaction between human hemoglobin (Hb) and oxali-palladium was studied using different spectroscopic methods of UV–vis, fluorescence, circular dichroism (CD), and chemiluminescence at two temperatures of 25 and 37°C. The experimental results showed that both dynamic and static quenching is occurred simultaneously when oxali-palladium quenches the fluorescence of Hb. According to the fluorescence quenching method, the binding site number, apparent binding constant, and corresponding thermodynamic parameters were measured at two temperatures. The values of ΔH°, ΔS°, and ΔG° indicate that process of the formation of oxali-palladium–Hb complex is a spontaneous interaction procedure in which electrostatic interaction plays a major role. In addition, UV–vis and CD results showed that the addition of oxali-palladium changes the conformation of Hb. To evaluate the functional changes of Hb via destruction of the heme structure, fluorescence studies were performed. The results demonstrated that two fluorescent heme degradation products are found during the interaction of oxali-palladium with Hb. Also, the amount of hydrogen peroxide produced in the solution of Hb due to the interaction of oxali-palladium with Hb using chemiluminescence method indicated heme degradation in the protein is occurred. Structural and functional changes induced in Hb via heme degradation are considered as side effects of this synthesized anticancer drug.     

Activation of c-Jun N-terminal kinase and apoptosis in endothelial cells mediated by endogenous generation of hydrogen peroxide

Reactive oxygen species have been implicated in the activation of signal transduction pathways. However, extracellular addition of oxidants such as hydrogen peroxide (H2O2) often requires concentrations that cannot be readily achieved under physiological conditions to activate biological responses such as apoptosis. Explanations for this discrepancy have included increased metabolism of H2O2 in the extracellular environment and compartmentalization within the cell. We have addressed this issue experimentally by examining the induction of apoptosis of endothelial cells induced by exogenous addition of H2O2 and by a redox cycling agent, 2,3-dimethoxy-1,4-naphthoquinone, that generates H2O2 in cells. Here we show that low nanomolar steady-state concentrations (0.1-0.5 nmol x min(-1) x 10(6) cells) of H2O2 generated intracellularly activate c-Jun N terminal kinase and initiate apoptosis in endothelial cells. A comparison with bolus hydrogen peroxide suggests that the low rate of intracellular formation of this reactive oxygen species results in a similar profile of activation for both c-Jun N terminal kinase and the initiation of apoptosis. However, a detailed analysis reveals important differences in both the duration and profile for activation of these signaling pathways.     

The effect of endogenous hydrogen peroxide induced by cold treatment in the improvement of tissue regeneration efficiency

We propose that oxidative stress resulting from an imbalance between generation and scavenging hydrogen peroxide contributes to tissue regeneration efficiency during somatic embryogenesis of hexaploid winter wheat (Triticum aestivum cv. Kamila) and organogenesis of faba bean (Vicia faba ssp. minor cv. Nadwislanski). Endogenous hydrogen peroxide content and antioxidant capacity of cells were determined in initial explants and callus cultures derived from these explants. Regeneration-competent explants (immature embryos) contained more endogenous H₂O₂ than explants initiated from regeneration-recalcitrant tissue (mature wheat embryos and faba bean epicotyls). Higher H₂O₂ levels were observed despite the higher activity of antioxidative enzymes (superoxide dismutase and catalase) and the induction of their gene expression. Calli originating from immature embryos retained the capacity of the initial explants: high H₂O₂ production was observed during the whole culture period. Low temperature treatment (4°C) was found to be an effective factor, which improved both regeneration ability and H₂O₂ production. Exogenous application to the medium of H₂O₂ and catalase blocker (3-aminotriazole), but not FeEDTA and superoxide dismutase blocker (diethyldithiocarbamate), also resulted in the enhancement of regeneration efficiency. These results clearly indicate that plant regeneration is specifically regulated by endogenous H₂O₂ and by factors, which improve its accumulation. Moreover, a study of the activity of various SOD isoforms suggests that not only the absolute concentration of H₂O₂, but also its localisation might be responsible for controlling regeneration processes.     

Effects of diethyldithiocarbamate and endogenous polyamine content on cellular responses to hydrogen peroxide cytotoxicity.

In exponential-phase Chinese-hamster cells, 0.1 mM-diethyldithiocarbamate (DDC) afforded greater than 1 log survival protection to cultures treated before and during exposure to 1 mM-H2O2. Both DDC and H2O2 treatment stimulated the activity of ornithine decarboxylase (ODC), the first enzyme in polyamine synthesis, within 4 h of exposure. DDC, and to a lesser degree H2O2, also stimulated the activity of spermidine N1-acetyltransferase (SAT), the rate-limiting enzyme in polyamine catabolism. The increase in SAT activity, after exposure to DDC or another stress (heat shock), was inhibited in cells depleted of putrescine and spermidine by alpha-difluoromethylornithine (DFMO), the enzyme-activated suicide inhibitor of ODC. Pretreatment with DFMO or heat shock also induced resistance to H2O2 cytotoxicity. Since SAT activity is low in resting cells, yet stimulation of enzyme activity depends on endogenous spermidine pools, these results suggest that the expression of SAT activity occurs by a mechanism involving a stress-dependent displacement of spermidine into a new intracellular compartment. The stimulation of ODC and SAT activities does not appear to be a necessary component of the mechanism by which DDC protects cells from H2O2 cytotoxicity, although spermidine displacement may be a common facet of the cellular response to stress.