Cariogenic Streptococcus mutans encounters a variety of host defense factors produced in oral cavity. Nitric oxide (NO) and NO-mediated reactive nitrogen species are potential antimicrobials of innate immunity that can threaten the fitness of S. mutans in their ecological niches. Streptococcal strategies to detoxify cytotoxic NO, which allow S. mutans to persist in caries or other environments of the oral cavity, remain unknown. In this study, we directly measured NO consumption rates of S. mutans isolated in Korea. Surprisingly, all S. mutans strains were unable to consume exogenous NO efficiently, while an intracellular parasite Salmonella enterica serovar Typhimurium expressing the NO-metabolizing enzyme flavohemoglobin consumed most of the NO. This result suggested that S. mutans has alternative detoxification systems for tolerating NO-induced nitrosative stresses.
Immunity, Innate ; Korea* ; Mouth ; Nitric Oxide* ; Parasites ; Reactive Nitrogen Species ; Salmonella enterica ; Streptococcus mutans* ; Streptococcus*

Reactive oxygen species (ROS) and nitrogen species (RNS) are both important signaling molecules involved in pain transmission in the dorsal horn of the spinal cord. Xanthine oxidase (XO) is a well-known enzyme for the generation of superoxide anions (O₂˙⁻), while S-nitroso-N-acetyl-DL-penicillamine (SNAP) is a representative nitric oxide (NO) donor. In this study, we used patch clamp recording in spinal slices of rats to investigate the effects of O₂˙⁻ and NO on the excitability of substantia gelatinosa (SG) neurons. We also used confocal scanning laser microscopy to measure XO- and SNAP-induced ROS and RNS production in live slices. We observed that the ROS level increased during the perfusion of xanthine and xanthine oxidase (X/XO) compound and SNAP after the loading of 2',7'-dichlorofluorescin diacetate (H₂DCF-DA), which is an indicator of intracellular ROS and RNS. Application of ROS donors such as X/XO, β-nicotinamide adenine dinucleotide phosphate (NADPH), and 3-morpholinosydnomimine (SIN-1) induced a membrane depolarization and inward currents. SNAP, an RNS donor, also induced membrane depolarization and inward currents. X/XO-induced inward currents were significantly decreased by pretreatment with phenyl N-tert-butylnitrone (PBN; nonspecific ROS and RNS scavenger) and manganese(III) tetrakis(4-benzoic acid) porphyrin (MnTBAP; superoxide dismutase mimetics). Nitro-L-arginine methyl ester (NAME; NO scavenger) also slightly decreased X/XO-induced inward currents, suggesting that X/XO-induced responses can be involved in the generation of peroxynitrite (ONOO⁻). Our data suggest that elevated ROS, especially O₂˙⁻, NO and ONOO⁻, in the spinal cord can increase the excitability of the SG neurons related to pain transmission.
Adenine ; Animals ; Humans ; Membranes ; Microscopy, Confocal ; Neurons* ; Nitric Oxide ; Nitrogen* ; Perfusion ; Peroxynitrous Acid ; Rats ; Reactive Oxygen Species* ; Spinal Cord ; Spinal Cord Dorsal Horn ; Substantia Gelatinosa* ; Superoxide Dismutase ; Superoxides ; Tissue Donors ; Xanthine ; Xanthine Oxidase

A growing body of evidence suggests that hyperglycemia-induced oxidative stress plays an important role in diabetic complications, especially beta-cell dysfunction and failure. Under physiological conditions, reactive oxygen species serve as second messengers that facilitate signal transduction and gene expression in pancreatic beta-cells. However, under pathological conditions, an imbalance in redox homeostasis leads to aberrant tissue damage and beta-cell death due to a lack of antioxidant defense systems. Taking into account the vulnerability of islets to oxidative damage, induction of endogenous antioxidant enzymes or exogenous antioxidant administration has been proposed as a way to protect beta-cells against diabetic insults. Here, we consider recent insights into how the redox response becomes deregulated under diabetic conditions, as well as the therapeutic benefits of antioxidants, which may provide clues for developing strategies aimed at the treatment or prevention of diabetes associated with beta-cell failure.
Antioxidants ; Choristoma ; Diabetes Complications ; Gene Expression ; Homeostasis ; Nitric Oxide ; Oxidation-Reduction ; Oxidative Stress ; Reactive Nitrogen Species ; Reactive Oxygen Species ; Second Messenger Systems ; Signal Transduction ; Superoxide Dismutase

PURPOSE: To evaluate the roles of macrophages and their influences on the signal transduction in the periprosthetic osteolysis, the pro-inflammatory signals were analyzed in particles-stimulated macrophages. MATERIALS AND METHODS: Raw 264.7 cell line derived from mice macrophages was used as pre-osteoclasts. To increase the stimulatory effects, the particles were composed of PMMA and polyethylene. Under the similar conditions as osteoclast differentiation, we examined the effect of particles on the pro-inflammatory signals in macrophage: the production of TNF-alpha, the activity of MAPKs (mitogen-activated phosphorylation kinase), the expression of I-kappaB (Inhibitory (B) and the production of H2O2 and nitric oxide. RESULTS: The particles stimulated the secretion of TNF-alpha and increased the phosphorylation of p38 and ERK in course of time. The concentration of H2O2 was increased; however the nitric oxide formation was not increased by particle treatment. In addition, the production of H2O2 was synergistically increased by suboptimal stimulation with PMA (phorbol 12-myristate 13-acetate). Expressions of I-kappaB were inhibited by particles. CONCLUSION: The particles may stimulate the activation of MAPKs, the production of TNF-alpha, reactive oxygen species (ROS) and the activation of NF-kappaB in the pre-osteoclasts. We speculate that particles may mediate the pro-inflammatory signal cascade by the activation of NF-kappaB through ROS in pre-osteoclasts, rather than reactive nitrogen species (RNS). Therefore we suggest that the macrophages in inflammatory osteolysis may have the characteristics of pre-osteoclast as well as pro-inflammatory cell. Further researches should be recommended.
Animals ; Cell Line ; Macrophages ; Mice ; NF-kappa B ; Nitric Oxide ; Osteoclasts ; Osteolysis ; Phosphorylation ; Polyethylene ; Polymethyl Methacrylate ; Reactive Nitrogen Species ; Reactive Oxygen Species ; Signal Transduction ; Tumor Necrosis Factor-alpha

Antimicrobial actions of reactive oxygen/nitrogen species (ROS/RNS) derived from products of NADPH oxidase and inducible nitric oxide (NO) synthase in host phagocytes inactivate various bacterial macromolecules. To cope with these cytotoxic radicals, pathogenic bacteria have evolved to conserve systems necessary for detoxifying ROS/RNS and repairing damages caused by their actions. In response to these stresses, bacteria also induce expression of molecular chaperones to aid in ameliorating protein misfolding. In this study, we explored the function of a newly identified chaperone Spy, that is localized exclusively in the periplasm when bacteria exposed to conditions causing spheroplast formation, in the resistance of Salmonella Typhimurium to ROS/RNS. A spy deletion mutant was constructed in S. Typhimurium by a PCR-mediated method of one-step gene inactivation with lambda Red recombinase, and subjected to ROS/RNS stresses. The spy mutant Salmonella showed a modest decrease in growth rate in NO-producing cultures, and no detectable difference of growth rate in H2O2 containing cultures, compared with that of wild type Salmonella. Quantitative RT-PCR analysis showed that spy mRNA levels were similar regardless of both stresses, but were increased considerably in Salmonella mutants lacking the flavohemoglobin Hmp, which are incapable of NO detoxification, and lacking an alternative sigma factor RpoS, conferring hypersusceptibility to H2O2. Results demonstrate that Spy expression can be induced under extreme conditions of both stresses, and suggest that the protein may have supportive roles in maintaining proteostasis in the periplasm where various chaperones may act in concert with Spy, thereby protecting bacteria against toxicities of ROS/RNS.
Bacteria ; Gene Silencing ; Molecular Chaperones ; NADPH Oxidase ; Nitric Oxide ; Periplasm ; Phagocytes ; Reactive Nitrogen Species ; Reactive Oxygen Species ; Recombinases ; RNA, Messenger ; Salmonella typhimurium ; Salmonella* ; Sigma Factor ; Spheroplasts

BACKGROUND:The present study was performed to further improve our understanding of the molecular mechanisms involved in the activation of NFkB, a major transcriptional factor involved in the inflammatory response in the inflammatory response in the lung, by particulate matter in lung epithelial cells wit an aerodynamic diameter of less than 2.5 micro meter(PM2.5). METHODS: Immediate production of reactive oxygen species (ROS) and nitrogen species (RNS), with the PM2.5 induced expression of inducible nitric oxide synthase (iNOS), IkB degradatio and NFkB-dependent transcrptional activity, in A 549 cells, were monitored. Addition, we also examined the effect of the iNOS inhibitor, L-N6-(1-iminoethyl) lysine hydrochloride (L-NIL), on the PM 2.5-induced NFkB activation in A 549 cells. RESULTS:The rapid degradation of IkB and the increase of transcriptional activity of the NFkB-dependent promotor were observed in A 549 cells exposed to PM2.5. The immediate production of ROS in response to PM2.5 in A 549 cells was not clearly detected, although immediate responses were observed in RAW 264.7 cells. A549 cells, cultured in the presence of PM2.5, produced an increase in NO, which was noticeably significant after 15 min of exposure with the expression of iNOS mRNA. The addition of L-NIL, an iNOS inhibitor, significantly inhibited the PM2.5-induced IkB degradation and the increase of the NFkB-dependent transcriptional activity. CONCLUSION: These results suggest that PM2.5 stimulates the immediate production of RNS, leading to the activation of NFkB in the pulmonary epithelium.
Cells, Cultured ; Epithelial Cells* ; Epithelium ; Lung* ; Lysine ; Nitric Oxide Synthase Type II ; Nitrogen ; Particulate Matter ; Reactive Oxygen Species ; RNA, Messenger

BACKGROUND:The present study was performed to further improve our understanding of the molecular mechanisms involved in the activation of NFkB, a major transcriptional factor involved in the inflammatory response in the inflammatory response in the lung, by particulate matter in lung epithelial cells wit an aerodynamic diameter of less than 2.5 micro meter(PM2.5). METHODS: Immediate production of reactive oxygen species (ROS) and nitrogen species (RNS), with the PM2.5 induced expression of inducible nitric oxide synthase (iNOS), IkB degradatio and NFkB-dependent transcrptional activity, in A 549 cells, were monitored. Addition, we also examined the effect of the iNOS inhibitor, L-N6-(1-iminoethyl) lysine hydrochloride (L-NIL), on the PM 2.5-induced NFkB activation in A 549 cells. RESULTS:The rapid degradation of IkB and the increase of transcriptional activity of the NFkB-dependent promotor were observed in A 549 cells exposed to PM2.5. The immediate production of ROS in response to PM2.5 in A 549 cells was not clearly detected, although immediate responses were observed in RAW 264.7 cells. A549 cells, cultured in the presence of PM2.5, produced an increase in NO, which was noticeably significant after 15 min of exposure with the expression of iNOS mRNA. The addition of L-NIL, an iNOS inhibitor, significantly inhibited the PM2.5-induced IkB degradation and the increase of the NFkB-dependent transcriptional activity. CONCLUSION: These results suggest that PM2.5 stimulates the immediate production of RNS, leading to the activation of NFkB in the pulmonary epithelium.
Cells, Cultured ; Epithelial Cells* ; Epithelium ; Lung* ; Lysine ; Nitric Oxide Synthase Type II ; Nitrogen ; Particulate Matter ; Reactive Oxygen Species ; RNA, Messenger

In this study, we investigated the antioxidative and antimicrobial activities of red algae Gloiopeltis tenax (GT). GT was extracted with methanol and then further fractionated it into four different types: methanol (GTMM), hexane (GTMH), butanol (GTMB) and aqueous (GTMA) soluble fractions. The antioxidant activity of the fractions from GT was investigated by measuring the scavenging activities of GT against reactive oxygen species (ROS) and reactive nitrogen species (RNS). Among the four fractions of GT, GTMM and GTMB showed a marked scavenging effect against ROS, but they displayed very low levels of the scavenging effect against RNS. The antimicrobial activity was increased in proportion to its concentration by the paper disc method. Among the various solvent layers, the GTMM and GTMB showed strong antimicrobial activities.
Methanol ; Reactive Nitrogen Species ; Reactive Oxygen Species ; Rhodophyta

Reactive oxygen species (ROS) and nitrogen species (RNS) are implicated in cellular signaling processes and as a cause of oxidative stress. Recent studies indicate that ROS and RNS are important signaling molecules involved in nociceptive transmission. Xanthine oxidase (XO) system is a well-known system for superoxide anions (O2(.-)) generation, and sodium nitroprusside (SNP) is a representative nitric oxide (NO) donor. Patch clamp recording in spinal slices was used to investigate the role of O2(.-) and NO on substantia gelatinosa (SG) neuronal excitability. Application of xanthine and xanthine oxidase (X/XO) compound induced membrane depolarization. Low concentration SNP (10 microM) induced depolarization of the membrane, whereas high concentration SNP (1 mM) evoked membrane hyperpolarization. These responses were significantly decreased by pretreatment with phenyl N-tert-butylnitrone (PBN; nonspecific ROS and RNS scavenger). Addition of thapsigargin to an external calcium free solution for blocking synaptic transmission, led to significantly decreased X/XO-induced responses. Additionally, X/XO and SNP-induced responses were unchanged in the presence of intracellular applied PBN, indicative of the involvement of presynaptic action. Inclusion of GDP-beta-S or suramin (G protein inhibitors) in the patch pipette decreased SNP-induced responses, whereas it failed to decrease X/XO-induced responses. Pretreatment with n-ethylmaleimide (NEM; thiol-alkylating agent) decreased the effects of SNP, suggesting that these responses were mediated by direct oxidation of channel protein, whereas X/XO-induced responses were unchanged. These data suggested that ROS and RNS play distinct roles in the regulation of the membrane excitability of SG neurons related to the pain transmission.
Animals ; Calcium ; Ethylmaleimide ; Humans ; Membranes ; Neurons* ; Nitric Oxide ; Nitrogen* ; Nitroprusside ; Oxidative Stress ; Rats* ; Reactive Oxygen Species* ; Substantia Gelatinosa* ; Superoxides ; Suramin ; Synaptic Transmission ; Thapsigargin ; Tissue Donors ; Xanthine ; Xanthine Oxidase

PURPOSE: The aim of this work was to investigate the beneficial effects of rhamnazin against inflammation, reactive oxygen species (ROS)/reactive nitrogen species (RNS), and anti-oxidative activity in murine macrophage RAW264.7 cells. METHODS: To examine the beneficial properties of rhamnazin on inflammation, ROS/ RNS, and anti-oxidative activity in the murine macrophage RAW264.7 cell model, several key markers, including COX and 5-LO activities, NO•, ONOO-, total reactive species formation, lipid peroxidation, •O₂ levels, and catalase activity were estimated. RESULTS: Results show that rhamnazin was protective against LPS-induced cytotoxicity in macrophage cells. The underlying action of rhamnazin might be through modulation of ROS/RNS and anti-oxidative activity through regulation of total reactive species production, lipid peroxidation, catalase activity, and •O₂, NO•, and ONOO• levels. In addition, rhamnazin down-regulated the activities of pro-inflammatory COX and 5-LO. CONCLUSION: The plausible action by which rhamnazin renders its protective effects in macrophage cells is likely due to its capability to regulate LPS-induced inflammation, ROS/ RNS, and anti-oxidative activity.
Arachidonate 5-Lipoxygenase ; Catalase ; Inflammation* ; Lipid Peroxidation ; Macrophages* ; Nitrogen ; Reactive Nitrogen Species ; Reactive Oxygen Species