Reactive oxygen intermediates modulate nitric oxide signaling in the plant hypersensitive disease-resistance response

The mechanisms involved in plant defense show several similar characteristics with the innate immune systems of vertebrates and invertebrates. In animals, nitric oxide (NO) cooperates with reactive oxygen intermediates (ROI) to kill tumor cells and is also required for macrophage killing of bacteria. Such cytotoxic events occur because unregulated levels of NO determine its diffusion-limited reaction with O₂^– generating peroxynitrite (ONOO^–), a mediator of cellular injury in many biological systems. In soybean suspension cells, unregulated NO production during the onset of a pathogen-induced hypersensitive response (HR) is not sufficient to activate the hypersensitive cell death, which is triggered only by fine tuning the NO/ROI ratio. Furthermore, that hypersensitive cell death is activated following interaction of NO with H₂O₂, rather than O₂^–. Increasing O₂^– levels reduces NO-derived toxicity, and the addition of ONOO^– to soybean suspensions does not affect cell viability. Consistently with the fact that ONOO^– is not an essential mediator of NO/RO-induced cell death, during the HR superoxide dismutase (SOD) accelerates O₂^– dismutation to H₂O₂ and therefore minimizes the loss of NO by reaction with O₂^– and triggers hypersensitive cell death through the NO/H₂O₂ synergism. Consequently, the rates of production and dismutation of O₂^– generated during the oxidative burst play a crucial role in modulating NO signaling through the cell death pathway, which proceeds through mechanisms different from those commonly observed in animals.     

NaCl胁迫下交替呼吸途径对烟草悬浮细胞死亡调节作用的研究

盐分胁迫是植物在自然环境中经常遭遇的环境胁迫因素之一,会引起植物代谢紊乱乃至细胞死亡,这严重限制了植物的生长、繁育和生存。交替呼吸途径是植物较之动物独特的线粒体呼吸途径。该研究在烟草悬浮细胞中调查了交替呼吸途径对Na Cl胁迫引起的植物细胞死亡过程的调节作用及相应的内在机制,以及在200 mmol·L~(-1)Na Cl处理的烟草悬浮细胞中研究了交替呼吸途径和细胞死亡发生及H_2O_2之间的关系。结果表明:(1)随着Na Cl处理浓度的增加,烟草悬浮细胞死亡水平逐渐增加,而交替呼吸途径的容量也逐渐上升。(2)与Na Cl处理相似,外源H_2O_2的处理也能导致烟草悬浮细胞死亡水平的增加。200 mmol·L~(-1)Na Cl的胁迫导致明显的细胞死亡发生和H_2O_2产量的显著性增加;而较之200 mmol·L~(-1)Na Cl胁迫下的细胞,用水杨基氧肟酸(交替呼吸途径的抑制剂)预处理后的细胞再置于200 mmol·L~(-1)Na Cl的胁迫下导致更高水平的细胞死亡和H_2O_2的产生。综上表明,高盐胁迫诱导了烟草悬浮细胞的交替呼吸途径的增加,而交替呼吸途径则可能通过抑制活性氧的产生而起到缓解细胞死亡发生的作用。     

In vivo and in situ visualization of early physiological events induced by heavy metals in pea root meristem

Heavy metals (HMs) are toxic pollutants, which can negatively affect the physiological processes of plants; moreover, HMs can be present in the food chain endangering people’s health. The aim of this study was to investigate the early physiological events during HM exposure in the root tips of the food plant Pisum sativum L. Ten-day-old pea plants were treated with 100 μM CdCl₂ or CuSO₄, in nutrient solution for 48 h. We studied the rapid formation of different reactive oxygen species (hydrogen peroxide H₂O₂ and superoxide radical O₂^·−) and reactive nitrogen species (nitric oxide NO· and peroxynitrite ONOO⁻) together with membrane damage and cell death in the meristem cells of pea roots using in vivo and in situ microscopic methods. In our experimental system, copper and cadmium induced the formation of H₂O₂ and NO. Two hours of heavy metal treatments resulted in an increased O₂^·− formation; however, later the level of this reactive molecule dramatically decreased. We found that high levels of NO were needed for ONOO⁻ production under HM exposure. A fast loss of membrane integrity and decreased cell viability were detected in root tips of copper-treated plants. The effects of cadmium seemed to be slower compared to copper, but this non-essential metal also caused cell death. We concluded that viability decreased when NO and H₂O₂ levels were simultaneously high in the same tissues. Using the NO scavenger it was also evidenced that NO generation is essential for cell death induction under copper or cadmium stress.     

Compensatory activation of ERK1/2 in Atg5-deficient mouse embryo fibroblasts suppresses oxidative stress-induced cell death

Despite of the increasing evidence that oxidative stress may induce non-apoptotic cell death or autophagic cell death, the mechanism of this process is unclear. Here, we report a role and a down-stream molecular event of Atg5 during oxidative stress-induced cell death. Compared to wild type (WT) cells, Atg5-deficient mouse embryo fibroblasts (Atg5^-/- MEFs) and Atg5 knockdown HT22 neuronal cells were more resistant to cell death induced by H₂O₂. On the contrary, Atg5^-/- MEFs were as sensitive to tumor necrosis factor (TNF)-α and cycloheximide as WT cells, and were more sensitive to cell death triggered by amino acid-deprivation than WT MEFs. Treatment with H₂O₂ induced the recruitment of a GFP-LC3 fusion protein and conversion of LC3 I to LC3 II, correlated with the extent of autophagosome formation in WT cells, but much less in Atg5-deficient cells. Among stress kinases, ERK1/2 was markedly activated in Atg5^-/- MEFs and Atg5 knockdown HT22 and SH-SY5Y neuronal cells. The inhibition of ERK1/2 by MEK1 inhibitor (PD98059) or dominant negative ERK2 enhanced the susceptibility of Atg5^-/- MEFs to H₂O₂-induced cell death. Further, reconstitution of Atg5 sensitized Atg5^-/- MEFs to H₂O₂ and suppressed the activation of ERK1/2. These results suggest that the inhibitory effect of Atg5 deficiency on cell death is attributable by the compensatory activation of ERK1/2 in Atg5^-/- MEFs during oxidative stress-induced cell death.     

Pyruvate secreted by human lymphoid cell lines protects cells from hydrogen peroxide mediated cell death

Reactive oxygen species (ROS) released from polymorphonuclear leukocytes and macrophages could cause DNA damage, but also induce cell death. Therefore inhibition of cell death must be an important issue for accumulation of genetic changes in lymphoid cells in inflammatory foci. Scavengers in the post culture medium of four lymphoid cell lines, lymphoblastoid cell lines (LCL), Raji, BJAB and Jurkat cells, were examined. Over 80% of cultured cells showed cell death 24 h after xanthine (X)/xanthine oxidase (XOD) treatment, which was suppressed by addition of post culture medium from four cell lines in a dose-dependent manner. H₂O₂ but not O^·-₂ produced by the X/XOD reaction was responsible for the cytotoxity, thus we used H₂O₂ as ROS stress thereafter. The H₂O₂-scavenging activity of post culture media from four cell lines increased rapidly at the first day and continued to increase in the following 2–3 days for LCL, Raji and BJAB cells. The scavenging substance was shown to be pyruvate, with various concentrations in the cultured medium among cell lines. Over 99% of total pyruvate was present in the extracellular media and less than 1% in cells. α-Cyano-4-hydroxycinnamate, a specific inhibitor of the H⁺-monocarbohydrate transporter, increased the H₂O₂-scavenging activity in the media from all four cell lines via inhibition of pyruvate re-uptake by cultured cells from the media. These findings suggest that lymphoid cells in inflammatory foci could survive even under ROS by producing pyruvate, so that accumulation of lymphoid cells with DNA damage is possible.     

Multi-mechanisms are involved in reactive oxygen species regulation of mTORC1 signaling

The mammalian target of rapamycin complex 1(mTORC1) integrates diverse signals to control cell growth, proliferation, survival, and metabolism. Role of reactive oxygen species (ROS) on mTORC1 signaling remains obscure and mechanisms through which ROS modulate mTORC1 are not known. We demonstrate that low doses ROS exposure stimulate mTORC1 while high concentrations or long-term ROS treatment decrease mTORC1 activity in vivo and in a variety of cell lines. The dose/time needed for inhibition or activation are cell type-dependent. In HEK293 cells hydrogen peroxide (H₂O₂) stimulates phosphorylation of AMP-activated kinase (AMPK) (T172) and Raptor (S792), enhances association of activated AMPK with Raptor. Furthermore, AMPK inhibitor compound c inhibits H₂O₂-induced Raptor (S792) phosphorylation and reverses H₂O₂-induced de-phosphorylation of mTORC1 downstream targets p70-S6K1 (T389), S6 (S235/236) and 4E-BP1 (T37/46). H₂O₂ also stimulates association of endogenous protein phosphatase 2A catalytic subunit (PP2Ac) with p70-S6K1. Like compound c, inhibitor of PP2A, okadaic acid partially reverses inactivation of mTORC1 substrates induced by H₂O₂. Moreover, inhibition of PP2A and AMPK partially rescued cells from H₂O₂-induced cell death. High doses of H₂O₂ inhibit while low doses of H₂O₂ activate mTORC1 both in TSC2^?/? P53^?/? and TSC2^+/+ P53^?/? MEFs. These data suggest that PP2A and AMPK-mediated phosphorylation of Raptor mediate H₂O₂-induced inhibition of mTORC1 signaling.     

Adiponectin and colon cancer: evidence for inhibitory effects on viability and migration of human colorectal cell lines

Exogenous hydrogen peroxide (H₂O₂) induces oxidative stress and apoptosis in cancer cells. This study evaluated the antiapoptotic effects of pan-caspase and caspase-3, -8, or -9 inhibitors on H₂O₂-treated Calu-6 and A549 lung cancer cells in relation to reactive oxygen species (ROS) and glutathione (GSH). Treatment with 50–500 μM H₂O₂ inhibited the growth of Calu-6 and A549 cells at 24 h and induced apoptosis in these cells. All the tested caspase inhibitors significantly prevented cell death in H₂O₂-treated lung cancer cells. H₂O₂ increased intracellular ROS levels, including that of O ₂ ^·? , at 1 and 24 h. It also increased the activity of catalase but decreased the activity of SOD. In addition, H₂O₂ triggered GSH deletion in Calu-6 and A549 cells at 24 h. It reduced GSH levels in Calu-6 cells at 1 h but increased them at 24 h. Caspase inhibitors decreased O ₂ ^·? levels in H₂O₂-treated Calu-6 cells at 1 h and these inhibitors decreased ROS levels, including that of O ₂ ^·? , in H₂O₂-treated A549 cells at 24 h. Caspase inhibitors partially attenuated GSH depletion in H₂O₂-treated A549 cells and increased GSH levels in these cells at 24 h. However, the inhibitors did not affect GSH deletion and levels in Calu-6 cells at 24 h. In conclusion, H₂O₂ induced caspase-dependent apoptosis in Calu-6 and A549 cells, which was accompanied by increases in ROS and GSH depletion. The antiapoptotic effects of caspase inhibitors were somewhat related to the suppression of H₂O₂-induced oxidative stress and GSH depletion.     

Dependence of leukemic cell proliferation and survival on H2O2 and l-arginine

The proliferation and/or survival of a variety of cells is dependent on cellular hydrogen peroxide (H₂O₂) production. We tested whether this was true of leukemic cells, using cell lines from leukemic patients (CEM, 697, Mn-60, and Tanoue). We found that addition of catalase inhibited proliferation of all cell lines and induced death in two. However, this turned out to be due to arginase contamination of the catalase. Pure arginase inhibited cell proliferation and survival, which was reversible by adding l-arginine, demonstrating the l-arginine dependency of these cells. The glutathione peroxidase mimetic ebselen killed the cells by a novel, rapid form of death, preceded by cell blebbing and prevented by N-acetylcysteine, suggesting toxicity is not due to ebselen's antioxidant activity. Addition of N-acetylcysteine to remove endogenous H₂O₂ stimulated survival and proliferation, suggesting that basal levels of H₂O₂ promoted cell death. Consistent with this, leukemic cell death was induced by adding as little as 5 μM H₂O₂. Ascorbic acid, even at 100 μM, induced death through H₂O₂ production. Thus H₂O₂ does not promote proliferation and survival, rather the opposite, and previous literature may have misinterpreted the effects of antioxidants. Arginase, H₂O₂, ascorbic acid, and ebselen might be useful in the treatment of leukemia.     

Role of reactive oxygen species in cardiac preconditioning: Study with photoactivated rose bengal in isolated rat hearts

Oxygen radical scavengers have been shown to prevent the development of ischemic preconditioning, suggesting that reactive oxygen species (ROS) might be involved in this phenomenon. In the present study, we have investigated whether direct exposure to ROS produced by photoactivated Rose Bengal (RB) could mimic the protective effects of ischemic preconditioning.

Methods In vitro generation of ROS from photoactivated RB in a physiological buffer was first characterised by ESR spectroscopy in the presence of 2,2,6,6-tetramethyl-1-piperidone (oxoTEMP) or 5,5-dimethyl-1-pyrroline-N-oxide (DMPO). In a second part of the study, isolated rat hearts were exposed for 2.5 min to photoactivated RB. After 5 min washout, hearts underwent 30 min no-flow normothermic ischemia followed by 30 min of reperfusion.

Results and Conclusions The production of singlet oxygen (¹O₂) by photoactivated RB in the perfusion medium was evidenced by the ESR detection of the nitroxyl radical oxoTEMPO. Histidine completely inhibited oxoTEMPO formation. In addition, the use of DMPO has indicated that (i) superoxide anions (O^·-₂) are produced directly and (ii) hydroxyl radicals (HO^·) are formed indirectly from the successive O^·-₂ dismutation and the Fenton reaction. In the perfusion experiments, myocardial post-ischemic recovery was dramatically impaired in hearts previously exposed to the ROS produced by RB photoactivation (¹O₂, O^·-₂, H₂O₂ and HO^·) as well as when ¹O₂ was removed by histidine (50 mM) addition. However, functional recovery was significantly improved when hearts were exposed to photoactivated RB in presence of superoxide dismutase (10⁵ IU/L) and catalase (10⁶ IU/L).

Further studies are now required to determine whether the cardioprotective effects of Rose Bengal in presence of O^·-₂ and H₂O₂ scavengers are due to singlet oxygen or to other species produced by Rose Bengal degradation.     

Effects of H2O2 on the growth, secretion, and metabolism of hybridoma cells in culture

Summary The effect of low concentrations of hydrogen peroxide (H₂O₂) (5 × 10⁻⁷−9.5 × 10⁻⁷ M) on cell growth and antibody production was investigated with murine hybridoma cells (Mark 3 and anti-hPL) in culture. Cell growth, measured by flow cytometry with morphological parameters, was significantly stimulated by H₂O₂ (8 × 10⁻⁷ M) but H₂O₂ concentration of 7 × 10⁻⁶ M and above increased cell death. H₂O₂ stimulation of antibody production was nonsignificant. The metabolism of cells treated with 8 × 10⁻⁷ or 1 × 10⁻⁵ M H₂O₂ was similar to that of the control in terms of glucose and glutamine consumption, lactate and ammonia production, and amino acid concentrations in the medium. The concentrations of lactate dehydrogenase, a marker of cell death, in test and control cells were similar. However, concentrations of intracellular free radicals measured by flow cytometry with dihydrorhodamine 123 (DHR 123) and dichlorofluorescein diacetate (DCFH-DA) as fluorochromes were different. The reactive oxygen species content of cells in 8 × 10⁻⁷ M H₂O₂ was similar to that of the controls, but there was a sudden, marked production of superoxide anions (detected with DHR 123) and H₂O₂ or peroxides (detected with DCFH-DA) by cells incubated with 1 × 10⁻⁵ M H₂O₂ which increased with increasing H₂O₂ until cell death.     

Repeated H2O2 exposure drives cell cycle progression in an in vitro model of ulcerative colitis

The production of hydrogen peroxide (H₂O₂) drives tumourigenesis in ulcerative colitis (UC). Recently, we showed that H₂O₂ activates DNA damage checkpoints in human colonic epithelial cells (HCEC) through c‐Jun N‐terminal Kinases (JNK) that induces p21^WAF1. Moreover, caspases circumvented the G1/S and intra‐S checkpoints, and cells accumulated in G2/M. The latter observation raised the question of whether repeated H₂O₂ exposures alter JNK activation, thereby promoting a direct passage of cells from G2/M arrest to driven cell cycle progression. Here, we report that increased proliferation of repeatedly H₂O₂‐exposed HCEC cells (C‐cell cultures) was associated with (i) increased phospho‐p46 JNK, (ii) decreased total JNK and phospho‐p54 JNK and (iii) p21^WAF1 down‐regulation. Altered JNK activation and p21^WAF1 down‐regulation were accompanied by defects in maintaining G2/M and mitotic spindle checkpoints through adaptation, as well as by apoptosis resistance following H₂O₂ exposure. This may cause increased proliferation of C‐cell cultures, a defining initiating feature in the inflammation‐carcinoma pathway in UC. We further suggest that dysregulated JNK activation is attributed to a non‐apoptotic function of caspases, causing checkpoint adaptation in C‐cell cultures. Additionally, loss of cell‐contact inhibition and the overcoming of senescence, hallmarks of cancer, contributed to increased proliferation. Furthermore, there was evidence that p54 JNK inactivation is responsible for loss of cell‐contact inhibition. We present a cellular model of UC and suggest a sinusoidal pattern of proliferation, which is triggered by H₂O₂‐induced reactive oxygen species generation, involving an interplay between JNK activation/inactivation, p21^WAF1, c‐Fos, c‐Jun/phospho‐c‐Jun, ATF2/phospho‐ATF2, β‐catenin/TCF4‐signalling, c‐Myc, CDK6 and Cyclin D2, leading to driven cell cycle progression.     

Photogeneration of singlet oxygen (1O2) and free radicals (Sen·-, O·-2) by tetra-brominated hypocrellin B derivative

To improve photodynamic activity of the parent hypocrellin B (HB), a tetra-brominated HB derivative (compound 1) was synthesized in high yield. Compared with HB, compound 1 has enhanced red absorption and high molar extinction coefficients. The photodynamic action of compound 1, especially the generation mechanism and efficiencies of active species (Sen^·-, O^·-₂ and ¹O₂) were studied using electron paramagnetic resonance (EPR) and spectrophotometric methods. In the deoxygenated DMSO solution of compound 1, the semiquinone anion radical of compound 1 is photogenerated via the self-electron transfer between the excited and ground state species. The presence of electron donor significantly promotes the reduction of compound 1. When oxygen is present, superoxide anion radical (O^·-₂) is formed via the electron transfer from Sens^·- to the ground state molecular oxygen. The efficiencies of Sens^·- and O^·-₂ generation by compound 1 are about three and two times as much as that of HB, respectively. Singlet oxygen (¹O₂) can be produced via the energy transfer from triplet compound 1 to ground state oxygen molecules. The quantum yield of singlet oxygen (¹O₂) is 0.54 in CHCl₃ similar to that of HB. Furthermore, it was found that the accumulation of Sens^·- would replace that of O^·-₂ or ¹O₂ with the depletion of oxygen in the sealed system.     

Ozone-Induced Cell Death Mediated with Oxidative and Calcium Signaling Pathways in Tobacco Bel-W3 and Bel-B Cell Suspension Cultures

Ozone (O₃)-induced cell death in two suspension-cultured cell lines of tobacco (Nicotiana tabacum L.) derived from Bel-W3 (hyper-sensitive to O₃) and Bel-B (highly tolerant to O₃) varieties were studied. By exposing the newly prepared cell lines to the pulse of ozonized air, we could reproduce the conditions demonstrating the difference in O₃ sensitivity as observed in their original plants, depending on the exposure time. Since O₃-induced acute cell death was observed in the dark, the requirement for photochemical reactions could be eliminated. Addition of several ROS scavengers and chelators inhibited the cell death induced by O₃, indicating that singlet oxygen (¹O₂), hydrogen peroxide (H₂O₂), hydroxyl radical and redox-active metals such as Fe²⁺ play central roles in O₃-induced acute damages to the cells. As expected, we observed the generation of ¹O₂ and H₂O₂ in the O₃-treated cells using chemiluminescent probes. On the other hand, an NADPH oxidase inhibitor, superoxide dismutase (SOD), and some SOD mimics showed no inhibitory effect. Thiols added as antioxidants unexpectedly behaved as prooxidants drastically enhancing the O₃-induced cell death. It is noteworthy that some ROS scavengers effectively rescued the cells from dying even treated after the pulse of O₃ exposure, confirming the post-ozone progress of ROS-dependent cell death mechanism. Since one of the key differences between Bel-B and Bel-W3 was suggested to be the capacity for ROS detoxification by catalase, the endogenous catalase activities were compared in vivo in two cell lines. As expected, catalase activity in Bel-B cells was ca. 7-fold greater than that in Bel-W3 cells. Interestingly, Ca²⁺ chelators added prior to (not after) the pulse of O₃ effectively inhibited the induction of cell death. In addition, increases in cytosolic Ca²⁺ concentration sensitive to Ca²⁺ chelators, ion channel blockers, and ROS scavengers were observed in the transgenic Bel-W3 cells expressing aequorin, suggesting the action of Ca²⁺ as a secondary messenger initiating the oxidative cell death. The O₃-induced calcium response in Bel-W3 cells was much greater than Bel-B cells. Based on the results, possible pathways for O₃-dependent generation of the lethal level of ROS and corresponding signaling mechanism for induction of cell death were discussed.Key Words: calcium, cell death, Nicotiana tabacum L., ozone, reactive oxygen species     

O2− from elicitor-induced oxidative burst is necessary for triggering phenylalanine ammonia-lyase activation and catharanthine synthesis in Catharanthus roseus cell cultures

Elicitor, derived from the cell walls of Aspergillus niger, induced rapid generation of reactive oxygen intermediates (ROI), including superoxide anion (O₂⁻) and hydrogen peroxide (H₂O₂), sequentially followed by phenylalanine ammonia-lyase (PAL) activation and catharanthine biosynthesis in Catharanthus roseus suspension cells. The elicitor-induced PAL activation and catharanthine biosynthesis were blocked by NAD(P)H oxidase inhibitor, diphenylene iodonium (DPI). O₂⁻ generated by the reaction of xanthine/xanthine oxidase (X/XO) triggered PAL activation and catharanthine biosynthesis of C. roseus cells in the absence of elicitor and reversed the inhibitory effect of DPI on elicitor-induced PAL activation and catharanthine biosynthesis. External application of H₂O₂ and catalase had no effect on PAL activity and catharanthine contents of C. roseus cells. The results demonstrated a causal relationship between elicitor-induced oxidative burst and PAL activation in C. roseus suspension cells and suggested a sequence of signaling events from ROI production to PAL activation and catharanthine synthesis. Within this sequence, O₂⁻ rather than H₂O₂ appeared to trigger the subsequent reactions.     

Visualization of Vascular Ca2+ Signaling Triggered by Paracrine Derived ROS

Oxidative stress has been implicated in a number of pathologic conditions including ischemia/reperfusion damage and sepsis. The concept of oxidative stress refers to the aberrant formation of ROS (reactive oxygen species), which include O₂^•-, H₂O₂, and hydroxyl radicals. Reactive oxygen species influences a multitude of cellular processes including signal transduction, cell proliferation and cell death^1-6. ROS have the potential to damage vascular and organ cells directly, and can initiate secondary chemical reactions and genetic alterations that ultimately result in an amplification of the initial ROS-mediated tissue damage. A key component of the amplification cascade that exacerbates irreversible tissue damage is the recruitment and activation of circulating inflammatory cells. During inflammation, inflammatory cells produce cytokines such as tumor necrosis factor-α (TNFα) and IL-1 that activate endothelial cells (EC) and epithelial cells and further augment the inflammatory response⁷. Vascular endothelial dysfunction is an established feature of acute inflammation. Macrophages contribute to endothelial dysfunction during inflammation by mechanisms that remain unclear. Activation of macrophages results in the extracellular release of O₂^•- and various pro-inflammatory cytokines, which triggers pathologic signaling in adjacent cells⁸. NADPH oxidases are the major and primary source of ROS in most of the cell types. Recently, it is shown by us and others^9,10 that ROS produced by NADPH oxidases induce the mitochondrial ROS production during many pathophysiological conditions. Hence measuring the mitochondrial ROS production is equally important in addition to measuring cytosolic ROS. Macrophages produce ROS by the flavoprotein enzyme NADPH oxidase which plays a primary role in inflammation. Once activated, phagocytic NADPH oxidase produces copious amounts of O₂^•- that are important in the host defense mechanism^11,12. Although paracrine-derived O₂^•- plays an important role in the pathogenesis of vascular diseases, visualization of paracrine ROS-induced intracellular signaling including Ca²⁺ mobilization is still hypothesis. We have developed a model in which activated macrophages are used as a source of O₂^•- to transduce a signal to adjacent endothelial cells. Using this model we demonstrate that macrophage-derived O₂^•- lead to calcium signaling in adjacent endothelial cells.     

Excess copper induces accumulation of hydrogen peroxide and increases lipid peroxidation and total activity of copper–zinc superoxide dismutase in roots of Elsholtzia haichowensis

The effects of excess copper (Cu) on the accumulation of hydrogen peroxide (H₂O₂) and antioxidant enzyme activities in roots of the Cu accumulator Elsholtzia haichowensis Sun were investigated. Copper at 100 and 300 μM significantly increased the concentrations of malondialdehyde and H₂O₂, and the activities of catalase (E.C. 1.11.1.6), ascorbate peroxidase (E.C. 1.11.1.11), guaiacol peroxidase (GPOD, E.C. 1.11.1.7) and superoxide dismutase (SOD, E.C. 1.15.1.1). Isoenzyme pattern and inhibitor studies showed that, among SOD isoforms, only copper–zinc superoxide dismutase (CuZn–SOD) increased. Excess Cu greatly increased the accumulation of superoxide anion (O₂ ^·−) and H₂O₂ in E. haichowensis roots. This study also provides the first cytochemical evidence of an accumulation of H₂O₂ in the root cell walls as a consequence of Cu treatments. Experiments with diphenyleneiodonium as an inhibitor of NADPH oxidase, 1,2-dihydroxybenzene-3,5-disulphonic acid as an O₂ ^·− scavenger, and N-N-diethyldithiocarbamate as an inhibitor of SOD showed that the source of H₂O₂ in the cell walls could partially be NADPH oxidase. The enzyme can use cytosolic NADPH to produce O₂ ^·−, which rapidly dismutates to H₂O₂ by SOD. Apoplastic GPOD and CuZn–SOD activities were induced in roots of E. haichowensis with 100 μM Cu suggesting that these two antioxidant enzymes may be responsible for H₂O₂ accumulation in the root apoplast.     

Reactive oxygen intermediates regulate cellular response to apoptotic stimuli: an hypothesis

Production of reactive oxygen intermediates (ROI) has been thought for a long time to adversely affect the physiology and survival of a cell. There is now a growing body of evidence to suggest that ROI such as superoxide anion (O^·-₂) and hydrogen peroxide (H₂O₂) can influence the growth, as well as death, of animal cells in vitro. The observation that cells release O^·-₂ or its dismutation product H₂O₂, either constitutively in the case of tumor cells or following cytokine stimulation, has led to the speculation that they might possibly serve as intercellular messengers to stimulate proliferation via mechanisms common to natural growth factors. However, as the balance between cell populations in an organism is tightly controlled by the rate of proliferation and death of constituent cells, an increase in cell numbers could reciprocally be viewed as deregulation of cell death. Hence, it is equally important to decipher how ROI influence the response of cells to signals that activate cell death pathway(s). We propose that ROI not only regulate proliferation but also affect cell sensitivity to triggers which activate the cellular suicide program (apoptosis) versus those that cause accidental (necrotic) cell death.     

Activation of an oxidative burst is a general feature of sensitive plants exposed to the air pollutant ozone

Ozone exposure stimulates an oxidative burst in leaves of sensitive plants, resulting in the generation and accumulation of hydrogen peroxide (H₂O₂) in tobacco and tomato, and superoxide (O₂^–?) together with H₂O₂ in Arabidopsis accessions. Accumulation of these reactive oxygen species (ROS) preceded the induction of cell death, and both responses co‐occurred spatially in the periveinal regions of the leaves. Re‐current ozone exposure of the sensitive tobacco cv. Bel W3 in closed chambers or in the field led to an enlargement of existing lesions by priming the border cells for H₂O₂ accumulation. Open top chamber experiments with native herbaceous plants in the field showed that Malva sylvestris L. accumulates O₂^–? at those sites that later exhibit plant cell death. Blocking of ROS accumulation markedly reduced ozone‐induced cell death in tomato, Arabidopsis and M. sylvestris. It is concluded that ozone triggers an in planta generation and accumulation of H₂O₂ and/or O₂^–? depending on the species, accession and cultivar, and that both these reactive oxygen species are involved in the induction of cell death in sensitive crop and native plants.     

两种外源化学物影响PEG胁迫对水稻根系的伤害

研究过氧化氢内源消除剂和交替氧化酶专一性抑制剂影响渗透胁迫对水稻根系的伤害。结果表明:PEG 6000胁迫抑制了水稻幼根的生长,降低了相对含水量、增加了H₂O₂含量,并导致细胞死亡。用5 mmol·L^-1二甲基硫脲(过氧化氢内源消除剂,dimethylthiourea,DMTU)预处理水稻幼根能明显降低PEG胁迫下水稻幼根过氧化氢的含量,缓解细胞死亡和相对含水量的降低,但对水稻根的生长影响较小。在PEG胁迫下,用1 mmol·L^-1水杨基氧肟酸(交替氧化酶专一性抑制剂,salicylhydroxamic acid,SHAM)预处理水稻幼根能显著降低水稻幼根的生长和相对含水量,并增加水稻幼根的过氧化氢含量和细胞的死亡程度。这说明DMTU能缓解PEG胁迫对水稻根系伤害,而SHAM加剧了PEG胁迫对水稻根系伤害。