Activation by nitric oxide of an oxidative-stress response that defends Escherichia coli against activated macrophages (original) (raw)
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Roles of nitric oxide in inducible resistance of Escherichia coli to activated murine macrophages
Infection and immunity, 1995
Nitric oxide (NO.) is produced as a cytotoxic free radical through enzymatic oxidation of L-arginine in activated macrophages. Pure NO. gas was previously found to induce the Escherichia coli soxRS oxidative stress regulon, which is readily monitored by using a soxS'::lac fusion. The soxRS system includes antioxidant defenses, such as a superoxide dismutase and a DNA repair enzyme for oxidative damage, and protects E. coli from the cytotoxicity of NO.-generating macrophages. Previous experiments involved exposing E. coli to a bolus of NO. rather than the steadily generated gas expected of activated macrophages. We show here detectable induction of soxS transcription by NO. delivered at rates as low as 25 microM/h. Maximal induction was observed at 25 microM NO. per h under anaerobic conditions but at 125 microM/h aerobically. After incubation with murine macrophages, soxS expression was induced in the phagocytosed bacteria up to approximately 30-fold after an 8-h exposure. This ...
Nitric Oxide Potentiates Hydrogen Peroxide-induced Killing of Escherichia coli
1995
Previously, we reported that nitric oxide (NO) provides significant protection to mammalian cells from the cytotoxic effects of hydrogen peroxide (H202). Murine neutrophils and activated macrophages, however, produce NO, H202, and other reactive oxygen species to kill microorganisms, which suggests a paradox. In this study, we treated bacteria (Escherichia coh~ with NO and H202 for 30 rain and found that exposure to NO resulted in minimal toxicity, but greatly potentiated (up to 1,000-fold) H202-mediated killing, as evaluated by a clonogenic assay. The combination of NO/H2O 2 induced DNA double strand breaks in the bacterial gehome, as shown by field-inverted gel electrophoresis, and this increased DNA damage may correlate with cell killing. NO was also shown to alter cellular respiration and decrease the concentration of the antioxidant glutathione to a residual level of 15-20% in bacterial cells. The iron chelator desferrioxamine did not stop the action of NO on respiration and glutathione decrease, yet it prevented the NO/H202 synergistic cytotoxicity, implicating metal ions as critical participants in the NO/H202 cytocidal mechanism. Our results suggest a possible mechanism of modulation of H202-mediated toxicity, and we propose a new key role in the antimicrobial macrophagic response for NO.
Nitric Oxide-Induced Nitrative Stress Involved in Microbial Pathogenesis
Journal of Pharmacological Sciences, 2005
The pathogenic mechanism of infections is a complicated but important scientific theme that is now attracting great attention because of its association with host-derived as well as microbial factors. Recent advances in free radical research revealed that reactive oxygen and nitrogen oxide species such as superoxide (O 2 −) and nitric oxide (NO) play a leading role in the pathogenesis of infections caused by viral pathogens including influenza virus and other RNA viruses. Although NO and O 2 − have antimicrobial activity against bacteria, fungi, and parasites, in some viral infections they have an opposite effect. This exacerbation caused by NO and O 2 − is mediated by reactive nitrogen oxides, for example, peroxynitrite (ONOO −), generated by reaction of NO with O 2 −. These nitrogen oxides have strong oxidation and nitration potential and can modify biological molecules, thereby creating oxidative and nitrative stress that contributes to pathogenic processes during viral infection. Nitrative stress-mediated 8-nitroguanosine formation during influenza or Sendai virus infection has been the focus of enormous interest because it involves unique biochemical and pharmacological properties such as redox activity and mutagenic potential. In this review, we discuss the nature and impact of nitrative stress in viral infection, with emphasis on nitrative stress-mediated viral pathogenesis, which we have recently been investigating.
Journal of Experimental Medicine, 2000
The contribution of the NADPH phagocyte oxidase (phox) and inducible nitric oxide (NO) synthase (iNOS) to the antimicrobial activity of macrophages for Salmonella typhimurium was studied by using peritoneal phagocytes from C57BL/6, congenic gp91 phox Ϫ / Ϫ , iNOS Ϫ / Ϫ , and doubly immunodeficient phox Ϫ / Ϫ iNOS Ϫ / Ϫ mice. The respiratory burst and NO radical (NO и ) made distinct contributions to the anti-Salmonella activity of macrophages. NADPH oxidasedependent killing is confined to the first few hours after phagocytosis, whereas iNOS contributes to both early and late phases of antibacterial activity. NO-derived species initially synergize with oxyradicals to kill S . typhimurium , and subsequently exert prolonged oxidase-independent bacteriostatic effects. Biochemical analyses show that early killing of Salmonella by macrophages coincides with an oxidative chemistry characterized by superoxide anion (O 2 и Ϫ ), hydrogen peroxide (H 2 O 2 ), and peroxynitrite (ONOO Ϫ ) production. However, immunofluorescence microscopy and killing assays using the scavenger uric acid suggest that peroxynitrite is not responsible for macrophage killing of wild-type S . typhimurium . Rapid oxidative bacterial killing is followed by a sustained period of nitrosative chemistry that limits bacterial growth. Interferon ␥ appears to augment antibacterial activity predominantly by enhancing NO и production, although a small iNOS-independent effect was also observed. These findings demonstrate that macrophages kill Salmonella in a dynamic process that changes over time and requires the generation of both reactive oxidative and nitrosative species.
Brazilian Journal of Medical and Biological Research
Sepsis, the leading cause of death in intensive care units, is associated with overproduction of nitric oxide (NO) due to inducible NO synthase (iNOS), responsible for some of the pathologic changes. Aminoguanidine (AG) is a selective iNOS inhibitor with reported inconsistent actions in sepsis. To investigate the influence of iNOS, we studied models of acute bacterial sepsis using acute challenges with aerobic (Escherichia coli) and anaerobic (Bacteroides fragilis) bacteria in the presence of AG. Six-week-old, 23 g, male and female BALB/c and C57Bl/6j mice, in equal proportions, were inoculated (ip) with bacteria in groups of 4 animals for each dose and each experiment in the absence or presence of AG (50 mg/kg, ip, starting 24 h before challenge and daily until day 6) and serum nitrate was measured by chemiluminescence. Both types of bacteria were lethal to mice, with an LD50 of 6 nephelometric units (U) for E. coli and 8 U for B. fragilis. Nitrate production peaked on the second d...
Nitrosative stress in Escherichia coli : reduction of nitric oxide
Biochemical Society Transactions, 2011
The ability of enteric bacteria to protect themselves against reactive nitrogen species generated by their own metabolism, or as part of the innate immune response, is critical to their survival. One important defence mechanism is their ability to reduce NO (nitric oxide) to harmless products. The highest rates of NO reduction by Escherichia coli K-12 were detected after anaerobic growth in the presence of nitrate. Four proteins have been implicated as catalysts of NO reduction: the cytoplasmic sirohaem-containing nitrite reductase, NirB; the periplasmic cytochrome c nitrite reductase, NrfA; the flavorubredoxin NorV and its associated oxidoreductase, NorW; and the flavohaemoglobin, Hmp. Single mutants defective in any one of these proteins and even the mutant defective in all four proteins reduced NO at the same rate as the parent. Clearly, therefore, there are mechanisms of NO reduction by enteric bacteria that remain to be characterized. Far from being minor pathways, the currentl...
Functional and Mechanistic Characterization of Bacterial Nitric Oxide Signaling Pathways
2016
Author(s): Rao, Minxi | Advisor(s): Marletta, Michael A. | Abstract: Nitric oxide (NO) is a well-established signaling molecule and cytotoxic agent in mammals. NO is synthesized by nitric oxide synthase (NOS) by macrophages at high concentrations as a key part of the host immune response, and at low concentrations in endothelial and neuronal cells as a signaling agent. In endothelial cells, the primary NO receptor is soluble guanylate cyclase (sGC), which contains a heme-nitric oxide/oxygen binding domain (H-NOX). Selective binding of NO to the H-NOX domain is responsible for activation of sGC. Thus, the mammalian NO signaling system involves NO synthesis by NOS, and NO sensing by the H-NOX domain of sGC. NOS and H-NOX proteins have also been identified in a number of bacterial species, including pathogens. Putative roles for bacterial NOS proteins include protection against oxidative stress and antibiotics, while bacterial H-NOX proteins have been shown to govern processes such as ...
Nitric oxide and bacteria-host interactions in Escherichia coli urinary tract infection
2008
ObjectivesTo examine whether urinary tract infection–associated stimuli could regulate heme oxygenase-1 (HO-1) expression and to asses the significance of HO-1 in protecting urinary tract epithelial cells against nitric oxide (NO)-induced damage.MethodsHeme oxygenase-1 expression was investigated in the human renal epithelial cell line A498 in response to the uropathogenic Escherichia coli (UPEC) strain IA2, the NO-donor DETA/NONOate (DETA/NO), and proinflammatory cytokines (interleukin-1β, tumor necrosis factor-α, and interferon-γ) using reverse transcriptase polymerase chain reaction and Western blot analysis. Cell viability was examined by the trypan blue exclusion test and light microscopy.ResultsThe HO-1 inducer hemin and DETA/NO increased HO-1 expression in A498 cells, and glutathione depletion further increased HO-1 expression in response to DETA/NO and hemin. Stimulation with a UPEC strain or cytokines did not upregulate HO-1 expression. The cytokines induced inducible NO sy...
The American Journal of Pathology, 2010
Escherichia coli K1 is a leading cause of neonatal meningitis in humans. In this study , we sought to determine the pathophysiologic relevance of inducible nitric oxide (iNOS) in experimental E. coli K1 meningitis. By using a newborn mouse model of meningitis , we demonstrate that E. coli infection triggered the expression of iNOS in the brains of mice. Additionally , iNOS ؊/؊ mice were resistant to E. coli K1 infection , displaying normal brain histology , no bacteremia , no disruption of the blood-brain barrier, and reduced inflammatory response. Treatment with an iNOS specific inhibitor , aminoguanidine (AG), of wild-type animals before infection prevented the development of bacteremia and the occurrence of meningitis. The infected animals treated with AG after the development of bacteremia also completely cleared the pathogen from circulation and prevented brain damage. Histopathological and micro-CT analysis of brains revealed significant damage in E. coli K1-infected mice , which was completely abrogated by AG administration. Peritoneal macrophages and polymorphonuclear leukocytes isolated from iNOS ؊/؊ mice or pretreated with AG demonstrated enhanced uptake and killing of the bacteria compared with macrophages and polymorphonuclear leukocytes from wild-type mice in which E. coli K1 survive and multiply. Thus , NO produced by iNOS may be beneficial for E. coli to survive inside the macrophages , and prevention of iNOS could be a therapeutic strategy to treat neonatal E. coli meningitis.