Resistance to Nitric Oxide-induced Necrosis in Heme Oxygenase-1 Overexpressing Pulmonary Epithelial Cells Associated with Decreased Lipid Peroxidation (original) (raw)
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Nitric oxide protects against cellular damage and cytotoxicity from reactive oxygen species
Proceedings of the National Academy of Sciences, 1993
Nitric oxide, NO, which is generated by various components of the immune system, has been presumed to be cytotoxic. However, NO has been proposed to be protective against cellular damage resulting during ischemia reperfusion. Along with NO there is often concomitant formation of superoxide/hydrogen peroxide, and hence a synergistic relationship between the cytotoxic effects of nitric oxide and these active oxygen species is frequently assumed. To study more carefully the potential synergy between NO and active oxygen species in mammalian cell cytotoxicity, we utilized either hypoxanthine/xanthine cell cytotoxicity, we utilized either hypoxanthine/xanthine oxidase (a system that generates superoxide/hydrogen peroxide) or hydrogen peroxide itself. NO generation was accomplished by the use of a class of compounds known as "NONOates," which release NO at ambient temperatures without the requirement of enzyme activation or biotransformation. When Chinese hamster lung fibroblast...
AJP: Lung Cellular and Molecular Physiology, 2008
Reactive species of oxygen and nitrogen have been collectively implicated in pulmonary oxygen toxicity, but the contributions of specific molecules are unknown. Therefore, we assessed the roles of several reactive species, particularly nitric oxide, in pulmonary injury by exposing wild-type mice and seven groups of genetically altered mice to >98% O 2 at 1, 3, or 4 atmospheres absolute. Genetically altered animals included knockouts lacking either neuronal nitric oxide synthase (nNOS −/− ), endothelial nitric oxide synthase (eNOS −/− ), inducible nitric oxide synthase (iNOS −/− ), extracellular superoxide dismutase (SOD3 −/− ), or glutathione peroxidase 1 (GPx1 −/− ), as well as two transgenic variants (S1179A and S1179D) having altered eNOS activities. We confirmed our earlier finding that normobaric hyperoxia (NBO 2 ) and hyperbaric hyperoxia (HBO 2 ) result in at least two distinct but overlapping patterns of pulmonary injury. Our new findings are that the role of nitric oxide in the pulmonary pathophysiology of hyperoxia depends both on the specific NOS isozyme that is its source and on the level of hyperoxia. Thus, iNOS predominates in the etiology of lung injury in NBO 2 , and SOD3 provides an important defense. But in HBO 2 , nNOS is a major contributor to pulmonary injury, whereas eNOS is protective. In addition, we demonstrated that nitric oxide derived from nNOS is involved in a neurogenic mechanism of HBO 2 -induced lung injury that is linked to central nervous system oxygen toxicity through adrenergic/cholinergic pathways.
Protection from the Dark Side of NO: Signaling and Cellular Defenses Against Nitric Oxide Toxicity
IUBMB Life (International Union of Biochemistry and Molecular Biology: Life), 2004
Although it is employed in biological systems for intercellular signaling or inflammatory responses, nitric oxide is not readily contained by cell membranes and so might damage surrounding nontarget cells. We have studied the genetic and biochemical basis of cellular resistance to the toxicity of NO. Inducible resistance mechanisms activate defense pathways that diminish lethal damage, prevent apoptosis, and may employ increased repair systems. A key inducible component of the cell response to NO toxicity is the enzyme heme oxygenase-1 (HO1). The activity of HO1 is necessary for the basic resistance of mammalian cells to NO-mediated cytotoxicity. However, the critical HO1-dependent reaction(s) responsible for NO resistance have not yet been identified. The induction of HO1 in response to NO depends on limited transcriptional activation and, in some cell types, on dramatic NOinduced stabilization of the HO1 mRNA. In human fibroblasts, this stabilization increases directly with the degree of NO exposure. Novel regulatory pathways appear to underlie the pathways of inducible NO resistance and NO-mediated mRNA stability.
British Journal of Pharmacology, 1996
The effects of oxygen free radical scavengers and endothelial cell-derived nitric oxide (EDNO) on the death of porcine cultured aortic endothelial cells exposed to exogenous superoxide-[xanthine (0.4 mM)/ xanthine oxidase (0.04 unit ml-')+ diethylenetriaminepentaacetic acid (DTPA, 10 uM)] or hydroxyl radical-generating system(s) [superoxide generating system+ ferric iron (Fe3, 0.1 mM) or peroxynitrite (0-100 uM)] have been evaluated. 2 Spin trapping studies using 5,5-dimethyl-l-pyrroline-N-oxide (DMPO) with electron paramagnetic resonance spectrometry were also conducted to determine qualitatively the oxidant species generated by the oxidant generating systems. 3 Endothelial cell injury provoked by the exogenous superoxide generating system was inhibited by catalase, DTPA and a hydroxyl radical scavenger (dimethyl sulphoxide, DMSO), but not by superoxide dismutase (SOD). Addition of Fe3+ to the superoxide generating system enhanced the cell injury. These suggested that the direct cytotoxicity of exogenous superoxide is limited, and that endogenous transition metal-dependent hydroxyl radical formation is involved in the cell injury. 4 An inhibitor of the constitutive NO-pathway, NG-monomethyl-L-arginine, did not influence cell injury induced by the superoxide generating system, suggesting that basal NO production is not responsible for the cytotoxicity. 5 Stimulation of endothelial cells with bradykinin enhanced cell injury provoked by the exogenous superoxide generating system, but not by the exogenous hydroxyl radical generating system. The enhancement by bradykinin was inhibited by NG-monomethyl-L-arginine and bradykinin B2-receptor antagonist, D-Arg-[Hyp3, Thi5'8, D-Phe7] bradykinin, suggesting that an interaction of NO with superoxide is involved in the enhanced cytotoxicity. A possible intermediate of this reaction, peroxynitrite, also caused endothelial cell injury in a concentration-dependent manner. 6 The modulatory effects of NO on hydroxyl radical-like activity (=formaldehyde production) from the superoxide generating system was also evaluated in a cell-free superoxide/NO generating system, consisting of xanthine/xanthine oxidase, DTPA, DMSO, and various amounts of a spontaneous NO generator, sodium nitroprusside (SNP) and were compared with those of Fe3". At doses up to 10 gM, SNP concentration-dependently increased the formaldehyde production while the higher concentrations of SNP decreased. The maximum amount of formaldehyde produced by SNP was 5 fold less than that produced by Fe3+ (0.1 mM). Peroxynitrite-induced formaldehyde formation was concentrationdependently inhibited by SNP. 7 We conclude that agonist-stimulated but not basal NO production acts as cytotoxic hydroxyl radical donor as well as the endogenous transition metal when endothelial cells are exposed to exogenous superoxide anion, while the modulatory effect of EDNO is limited by a secondary reaction with hydroxyl radicals.
Two faces of nitric oxide: implications for cellular mechanisms of oxygen toxicity
Journal of Applied Physiology, 2008
Recent investigations have elucidated some of the diverse roles played by reactive oxygen and nitrogen species in events that lead to oxygen toxicity and defend against it. The focus of this review is on toxic and protective mechanisms in hyperoxia that have been investigated in our laboratories, with an emphasis on interactions of nitric oxide (NO) with other endogenous chemical species and with different physiological systems. It is now emerging from these studies that the anatomical localization of NO release, which depends, in part, on whether the oxygen exposure is normobaric or hyperbaric, strongly influences whether toxicity emerges and what form it takes, for example, acute lung injury, central nervous system excitation, or both. Spatial effects also contribute to differences in the susceptibility of different cells in organs at risk from hyperoxia, especially in the brain and lungs. As additional nodes are identified in this interactive network of toxic and protective respo...
Cellular antioxidant and pro-oxidant actions of nitric oxide
Free Radical Biology and Medicine, 1999
We describe a biphasic action of nitric oxide (NO) in its effects on oxidative killing of isolated cells: low concentrations protect against oxidative killing, while higher doses enhance killing, and these two effects occur by distinct mechanisms. While low doses of NO (from (Z)-1-[N-(3-ammonio propyl)-N-(n-propyl)-amino]-diazen-1-ium-1,2 2 diolate [PAPA/NO] or S-nitroso-N-acetyl-L-penicillamine [SNAP] prevent killing of rat hepatocytes by t-butylhydroperoxide (tBH), further increasing doses result in increased killing. Similar effects occur with rat hepatoma cells treated with PAPA/NO and tBH or H 2 O 2. Increased killing with higher concentrations of NO donor is due to both NO and tBH, because NO donor alone is without effect. Glutathione (GSH) is not involved in either of these actions. Based on measurements of thiobarbituric acid-reactive substances (TBARS) and effects of lipid radical scavenger (DPPD) and deferoxamine, the protective effect, but not the enhancing effect, involves peroxidative chemistry. Fructose has no effect on tBH killing alone but provides substantial protection against killing by higher concentrations of NO plus tBH, suggesting that the enhancing effect involves mitochondrial dysfunction. Hepatocytes, when stimulated to produce NO endogenously, become resistant to tBH killing, indicative of the presence of an NO-triggered antioxidant defensive mechanism. The finding that the protective effects of low concentrations of NO and the harmful effects of high concentrations of NO are fundamentally different in nature suggest that therapeutic interventions could be designed, which selectively prevent its pro-oxidant activity at high concentrations, thus converting NO from a "Janus-faced" modulator of oxidant injury into a "pure" protectant.
Mechanisms of the Antioxidant Effects of Nitric Oxide
Antioxidants & Redox Signaling, 2001
The Janus face of nitric oxide (NO) has prompted a debate as to whether NO plays a deleterious or protective role in tissue injury. There are a number of reactive nitrogen oxide species, such as N 2 O 3 and ONOO 2 , that can alter critical cellular components under high local concentrations of NO. However, NO can also abate the oxidation chemistry mediated by reactive oxygen species such as H 2 O 2 and O 2 2 that occurs at physiological levels of NO. In addition to the antioxidant chemistry, NO protects against cell death mediated by H 2 O 2 , alkylhydroperoxides , and xanthine oxidase. The attenuation of metal/peroxide oxidative chemistry, as well as lipid peroxidation, appears to be the major chemical mechanisms by which NO may limit oxidative injury to mammalian cells. In addition to these chemical and biochemical properties, NO can modulate cellular and physiological processes to limit oxidative injury, limiting processes such as leukocyte adhesion. This review will address these aspects of the chemical biology of this multifaceted free radical and explore the beneficial effect of NO against oxidative stress. Antioxid. Redox Signal. 3, 203-213.
Biological Chemistry, 2000
Nitric oxide (NO) plus oxygen (O 2 ) are known to cause cell damage via formation of reactive nitrogen species. NO itself directly inhibits cytochrome oxidase of the mitochondrial respiratory chain in competition with O 2 , thus inducing a hypoxic-like injury. To assess the critical NO and O 2 concentrations for both mechanisms of NOinduced cell injury, cells of a rat liver sinusoidal endothelial cell line were incubated in the presence of the NO donor spermineNONOate at different O 2 concentrations, and their loss of viability was determined by the release of lactate dehydrogenase. Protection by ascorbic acid was used as indication for the involvement of reactive nitrogen species, whereas a hypoxic-like injury was indicated by the protective effects of glycine and glucose and the increase in NAD(P)H fluorescence. High concentrations of NO (approx. 10 mM NO) and O 2 (21% O 2 ) were required to induce endothelial cell death mediated by formation of reactive nitrogen species. On the other hand, pathophysiologically relevant NO concentrations at low but physiological O 2 concentrations (ca. 2 mM NO at 5% O 2 and about 1 mM NO at 2% O 2 ) induced hypoxic-like cell death in the endothelial cells that was prevented by the presence of glucose.
Nitric oxide and peroxynitrite-mediated pulmonary cell death
The American journal of physiology, 1998
Nitric oxide (.NO) can be produced within the lung, and recently inhaled nitric oxide has been used as a therapeutic agent. Peroxynitrite1 (ONOO-), the product of the nearly diffusion-limited reaction between .NO and superoxide, may represent the proximal reactive species mediating .NO injury to pulmonary cells. To investigate the physiological and pathological reactivities of .NO and ONOO- at the molecular and cellular levels, bovine pulmonary artery endothelial cells (BPAEC) and rat type II epithelial cells were exposed to .NO (0.01-2.5 microM/min for 2 h) generated by spermine-NONOate and papa-NONOate and to the same fluxes of ONOO- generated by 1,3-morpholinosydnonimine (SIN-1). Exposure to SIN-1 resulted in cellular injury and death in both cell types. Epithelial cells displayed a concentration-dependent loss of cellular viability within 8 h of exposure. In contrast, BPAEC loss of cellular viability was evident after 18 h postexposure. Events preceding cell death in BPAEC inclu...