Augmentation of cytokine-induced nitric oxide synthesis by hydrogen peroxide

1996 ◽  
Vol 271 (1) ◽  
pp. L114-L120 ◽  
Author(s):  
S. A. Milligan ◽  
M. W. Owens ◽  
M. B. Grisham
Keyword(s):  
Nitric Oxide ◽  
Interleukin 1 ◽  
Northern Blotting ◽  
Oxidation Products ◽  
Tnf Alpha ◽  
Inos Mrna ◽  
Transcriptional Level ◽  
Necrosis Factor Alpha ◽  
Steady State Levels ◽  
Stimulation Of

The inducible isoform of nitric oxide synthase (iNOS) is induced upon stimulation of cells with cytokines and lipopolysaccharide (LPS). Stimulation of rat pleural mesothelial cells with combinations of interleukin-1 beta (IL-1 beta), tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma), and LPS induced the synthesis of nitric oxide as measured by the oxidation products nitrite (NO2-) and nitrate (NO3-). Addition of 25-50 microM H2O2 to the cytokines significantly augmented the synthesis of NO2- and NO3-. Stimulation with IL-1 beta and TNF-alpha plus H2O2 or IL-1 beta and LPS plus H2O2 increased the synthesis of NO2- and NO3- by 3.8- and 3.5-fold, respectively. These effects were inhibited by NG-nitro-L-arginine methyl ester and cycloheximide as well as by catalase. Immunoblotting demonstrated that H2O2 augmented cytokine-induced synthesis of iNOS protein. These effects were inhibited by certain antioxidants and metal chelators, suggesting that the hydroxyl radical may mediate the oxidant-induced effect. Northern blotting demonstrated that H2O2 greatly augmented steady-state levels of iNOS mRNA, suggesting that H2O2 acted in part at the transcriptional level.

Regulation of inducible nitric oxide synthase in hepatic sinusoidal endothelial cells

1996 ◽  
Vol 271 (2) ◽  
pp. G260-G267 ◽  
Author(s):  
D. C. Rockey ◽  
J. J. Chung
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Interleukin 1 ◽  
Cell Types ◽  
Tnf Alpha ◽  
Inos Mrna ◽  
No Production ◽  
Sinusoidal Endothelial Cells ◽  
Necrosis Factor Alpha ◽  
Ifn Gamma

Nitric oxide (NO) has many important physiological effects that depend in part on its cellular source(s). In liver, NO is produced by all major cell types, including hepatocytes, Kupffer, stellate, and sinusoidal endothelial cells (SECs). Although endothelial cells have been commonly associated with constitutive NO production, recent evidence suggests that NO is inducible in this cell type. Here, we investigated the regulation of inducible NO synthase (iNOS) in SECs. Interferon-gamma (IFN-gamma) and lipopolysaccharide (LPS) as individual compounds induced iNOS mRNA in SECs. Interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF-alpha) had no effect when used alone but enhanced iNOS mRNA upregulation by IFN-gamma. iNOS transcription after LPS was present only for 4 h after exposure yet was more sustained after IFN-gamma/TNF-alpha, LPS was unique in that it transiently induced iNOS mRNA, whereas IFN-gamma/TNF-alpha resulted in prolonged increases in iNOS mRNA. Both LPS and IFN-gamma/TNF-alpha caused prolonged elevation of immunoreactive protein. However, when stimulated by LPS, iNOS remained enzymatically active for only 24-48 h. After IFN-gamma or IFN-gamma/TNF-alpha, iNOS activity declined only moderately. LPS added to IFN-gamma alone or IFN-gamma/TNF-alpha did not result in more rapid decay of iNOS enzymatic activity. These data indicate that induction of iNOS by sinusoidal endothelial cells is prominent and that it is regulated both transcriptionally and by its inactivation. Such complex regulation of iNOS has important implications for NO biology in liver disease.

scholarly journals Suppression of interleukin 8 production by progesterone in rabbit uterine cervix

1994 ◽  
Vol 301 (1) ◽  
pp. 183-186 ◽  
Author(s):  
A Ito ◽  
K Imada ◽  
T Sato ◽  
T Kubo ◽  
K Matsushima ◽  
...  
Keyword(s):  
Matrix Metalloproteinases ◽  
Interleukin 1 ◽  
Interleukin 8 ◽  
Cervical Ripening ◽  
Transcriptional Level ◽  
Dependent Manner ◽  
Necrosis Factor Alpha ◽  
Steady State Levels ◽  
Neutrophil Chemotactic Factor

Uterine cervical fibroblasts prepared from rabbits at 23 days of gestation were found to produce spontaneously the neutrophil chemotactic factor/interleukin 8 (IL-8). When the cells were treated with recombinant human interleukin 1 alpha and 1 beta (rhIL-1 alpha and -1 beta), both cytokines similarly enhanced the production of IL-8 in a dose-dependent manner. Recombinant tumour necrosis factor alpha also enhanced its production to a lesser extent, but interleukin 6 failed to modulate the production. Physiological concentrations of progesterone suppressed both the spontaneous and IL-1-mediated production of IL-8 in parallel with the decrease in the steady-state levels of its mRNA. These suppressive actions of progesterone were offset by co-treatment of cells with a progesterone antagonist, mifepristone (RU486). In conclusion, basal and IL-1-induced IL-8 production in rabbit uterine cervical fibroblasts is down-regulated by progesterone at the transcriptional level. These results obtained in vitro and our previous observations indicating that progesterone modulates the extra-cellular matrix breakdown via the suppression of production of matrix metalloproteinases and the augmentation of production matrix metalloproteinases and the augmentation of production of their specific inhibitors (TIMP-1) [Sato, Ito, Mori, Yamashita, Hayakawa and Nagase (1991) Biochem. J. 275, 645-650] may explain the mechanisms of the maintenance of pregnancy until parturition and the acceleration of uterine cervical ripening and dilatation at term.

Pulmonary alveolar epithelial inducible NO synthase gene expression: regulation by inflammatory mediators

1995 ◽  
Vol 268 (3) ◽  
pp. L501-L508 ◽  
Author(s):  
H. H. Gutierrez ◽  
B. R. Pitt ◽  
M. Schwarz ◽  
S. C. Watkins ◽  
C. Lowenstein ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Gene Expression Regulation ◽  
Interleukin 1 ◽  
Alveolar Epithelium ◽  
Cell Types ◽  
Oxidation Products ◽  
No Oxidation ◽  
Target Cells ◽  
Cell Permeabilization ◽  
Necrosis Factor Alpha

Nitric oxide (.NO) is a short-lived mediator that can be induced by different cytokines and lipopolysaccharide (LPS) in a variety of cell types and produces many physiological and metabolic changes in target cells. In the current study, we show that a combination of cytokines, LPS, and zymosan-activated serum (ZAS; called for convenience cytomix Z) induces production of high concentrations of the NO oxidation products nitrite (NO2-) and nitrate (NO3-) by cultured rat fetal lung epithelial type II cells in a time-dependent fashion. Interferon-gamma and tumor necrosis factor-alpha alone did not significantly affect .NO synthesis, whereas ZAS, LPS, and interleukin-1 beta caused only a modest increase in formation of .NO oxidation products. Production of NO2- and NO3- was inhibited by NG-monomethyl-L-arginine and cyclohexmide. After exposure of these cells to a combination of the above cytokines, Escherichia coli LPS, and ZAS (cytomix Z), enhanced inducible nitric oxide synthase (iNOS) expression was indicated by an elevation in steady-state mRNA specific for iNOS (via Northern blot analysis) and increased immunofluorescence for iNOS after cell permeabilization, incubation with anti-iNOS antibody, and treatment with Cy3.18-conjugated rabbit-specific antibody. The extent of inflammatory mediator-induced.NO production by alveolar epithelium, which exceeds that of other lung cell types, reveals new insight into mechanisms of pulmonary host defense and pathways of free radical-mediated lung injury.

Roles of IL-1 and TNF-alpha in endotoxin-induced activation of nitric oxide synthase in cultured rat brain cells

1996 ◽  
Vol 270 (2) ◽  
pp. R326-R332 ◽  
Author(s):  
L. I. Romero ◽  
J. B. Tatro ◽  
J. A. Field ◽  
S. Reichlin
Keyword(s):  
Nitric Oxide ◽  
Interleukin 1 ◽  
Primary Cultures ◽  
Neonatal Rat ◽  
Tnf Alpha ◽  
Brain Cells ◽  
Nitrite Accumulation ◽  
No Production ◽  
Necrosis Factor Alpha ◽  
Oxide Synthase

In astrocytes and microglia, bacterial lipopolysaccharide (LPS) stimulates production and release of interleukin-1 beta (IL-1 beta), tumor necrosis factor-alpha (TNF-alpha), and nitric oxide (NO). Although IL-1 beta and TNF-alpha are themselves capable of inducing NO synthase (NOS) in glia, the specific factors mediating LPS induction of NOS in brain have not been identified. To determine whether LPS induction of NOS in brain cells is mediated by IL-1 or TNF-alpha, acting alone or in concert, the effects of IL-1-receptor antagonist (IL-1Ra) and of TNF-soluble receptor (TNFsRp55), presented individually and in combination, on LPS-induced NOS activity were tested. In glial-enriched mixed primary cultures of neonatal rat telencephalic cells, LPS (0.1-100 ng/ml), IL-1 beta (0.01-10 nM), and TNF-alpha (0.1-100 nM) each concentration dependently stimulated accumulation of nitrite, an indicator of NO production. Induction of nitrite accumulation by LPS and by IL-1 was blocked by N omega-nitro-L-arginine methyl ester and N omega-monomethyl-L-arginine, indicating that it was mediated by NOS. TNF-alpha alone induced NO production weakly as compared with IL-1, but combined submaximal concentrations of IL-1 beta (1 nM) and TNF-alpha (10 nM) induced NOS synergistically. Furthermore, TNFsRp55 and IL-1Ra each produced a dose-dependent partial inhibition of the NO response to LPS, and the effect of TNFsRp55 was equal to or greater than that of IL-1Ra. TNFsRp55 and IL-1Ra in combination were not significantly more effective than TNF-sRp55 alone. The results indicate that LPS induction of NOS activity in brain cells is mediated in part by both IL-1 beta and TNF-alpha.

scholarly journals Intraperitoneal injection of tetracyclines protects mice from lethal endotoxemia downregulating inducible nitric oxide synthase in various organs and cytokine and nitrate secretion in blood.

1997 ◽  
Vol 41 (1) ◽  
pp. 117-121 ◽  
Author(s):  
S Milano ◽  
F Arcoleo ◽  
P D'Agostino ◽  
E Cillari
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Inducible Nitric Oxide Synthase ◽  
Intraperitoneal Injection ◽  
Interleukin 1 ◽  
Tnf Alpha ◽  
Necrosis Factor Alpha ◽  
Oxide Synthase ◽  
Inducible Nitric Oxide

We have tested whether tetracyclines (TETs) are able to protect mice from lipopolysaccharide (LPS)-induced shock, a cytokine-mediated inflammatory reaction. Mice, injected with a single dose of tetracycline base (TETb; 1.5, 10 and 20 mg/kg of body weight) or doxycycline (DOXY; 1.5 mg/kg), were significantly protected from a lethal intraperitoneal injection of LPS (500 micrograms per mouse). TETs acted in early events triggered in response to LSP; in fact, they were no longer significantly protective if injected more than 1 h after the injection of endotoxin. LPS-treated mice protected by TETs showed a significant inhibition of tumor necrosis factor alpha (TNF-alpha), interleukin-1 alpha (IL-1 alpha), and nitrate secretion in the blood, events that were directly related with the survival. In mice treated with TETs a significant decrease of inducible nitric oxide synthase (iNOS) activity was observed in spleen and peritoneal cells compared with that detected in mice treated with LPS alone. Furthermore, TETs were found to inhibit NO synthesis by peritoneal macrophages stimulated in vitro with LPS. On the contrary, TETs were unable to decrease the ability of the macrophages to synthesize IL-1 alpha and TNF-alpha in vitro. These results indicate that TETs are not able to act directly on the synthesis of these cytokines, but they may modulate other pathways that could in turn be responsible for the inhibition of IL-1 alpha and TNF-alpha synthesis. Altogether, these results indicate that TETs are advantageous candidates for the prophylaxis and treatment of septic shock in mice, having both antimicrobial activity and the ability to inhibit endogenous TNF-alpha, IL-1 alpha, and iNOS, hence, exerting, potent anti-inflammatory effects.

Dissociation of TNF-alpha from endotoxin-induced nitric oxide and acute-phase hypotension

1997 ◽  
Vol 273 (1) ◽  
pp. H164-H174 ◽  
Author(s):  
J. Xie ◽  
K. O. Joseph ◽  
G. J. Bagby ◽  
T. D. Giles ◽  
S. S. Greenberg
Keyword(s):  
Nitric Oxide ◽  
Acute Phase ◽  
Ex Vivo ◽  
Plasma Concentrations ◽  
Platelet Activating Factor ◽  
Polymorphonuclear Neutrophils ◽  
Tnf Alpha ◽  
Inos Mrna ◽  
No Production ◽  
Necrosis Factor Alpha

We tested the concept that tumor necrosis factor-alpha (TNF-alpha) or platelet-activating factor (PAF) mediated Escherichia coli endotoxin lipopolysaccharide (LPS)-induced upregulation of nitric oxide (NO) and acute-phase hypotension (APH) in the rat. LPS (0.5 mg/kg i.v.) given to rats treated with saline or nonimmune goat-derived gamma-globulin (immunoglobulin G, 22 mg/kg i.m.) produced APH and increased plasma concentrations of TNF-alpha and nitrate and nitrite anions (reactive nitrogen intermediates; RNI) and NO in ex vivo incubates of polymorphonuclear neutrophils (PMN) and inducible NO synthase (iNOS) mRNA in PMN. Pretreatment of rats with a polyclonal TNF-alpha antibody (TNF-Ab, 22 mg/kg i.m.) abolished LPS-mediated increases in plasma TNF-alpha but failed to inhibit APH or the NO system. TNF-alpha (8.2 micrograms/kg i.v.) produced transient hypertension and sustained tachycardia and increased plasma TNF-alpha and PMN iNOS mRNA but not RNI. LPS and TNF-alpha decreased spontaneous and calcimycin (Ca2+ ionophore, 1 microM)- and PAF (1 microM)-mediated increases in head-space NO production by rings of mesenteric artery incubated ex vivo. TNF-Ab abolished all effects of TNF-alpha. PAF (25, 50, and 100 ng/kg) produced APH without increasing plasma TNF-alpha, RNI, or PMN iNOS mRNA. The PAF receptor antagonist BN-50730 (80 micrograms/kg i.v.) abolished PAF-induced APH and attenuated LPS-induced increases in RNI. We conclude that 1) LPS produces parallel but unrelated changes in TNF-alpha and RNI in plasma and PMN during the APH of endotoxemia; and 2) endogenous TNF-alpha is not required for LPS-mediated induction of iNOS mRNA, and PAF mediates LPS-induced APH.

Human Peritoneal Mesothelial Cells Produce Nitric Oxide: Induction by Cytokines

2000 ◽  
Vol 20 (6) ◽  
pp. 772-777 ◽  
Author(s):  
Jinn-Yang Chen ◽  
Jen-Hwey Chiu ◽  
Hui-Ling Chen ◽  
Tzen-Wen Chen ◽  
Wu-Chang Yang ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Interleukin 1 ◽  
Mesothelial Cells ◽  
Inos Mrna ◽  
No Production ◽  
Necrosis Factor Alpha ◽  
Peritoneal Mesothelial Cells ◽  
Human Peritoneal Mesothelial Cells ◽  
The Media ◽  
Nitrate And Nitrite

Objective To investigate the induction of nitric oxide synthase type II (iNOS) in human peritoneal mesothelial cells (HPMC) using cytokines and bacterial lipopolysaccharide (LPS). Design Confluent monolayers of HPMC were exposed to cytokines [tumor necrosis factor alpha (TNFα), interleukin-1 beta (IL-1β), interferon gamma (IFNγ)] or LPS, individually or in various double and triple combinations, for 24 – 72 hours. Concentrations of nitrate and nitrite in the media were quantified using the Griess reaction and used as indirect indices of nitric oxide (NO) production. The expression of iNOS was assessed using reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blot. Results Neither single cytokines nor LPS was able to induce iNOS mRNA or NO production. Both double combinations of TNFα+ IFNγ and IL-1β + IFNγ were able to induce iNOS mRNA expression, but only TNFα + IFNγ induced significant NO production. The triple combination of TNFα + IFNγ + IL-1β induced even more NO production than TNFα + IFNγ. There was no constitutive NO synthase type III (eNOS) expression in HPMC. Conclusions Certain combinations of cytokines could stimulate cultured HPMC to produce NO, and HPMC might be a source of intraperitoneal NO production during peritonitis.

scholarly journals Inducible Nitric Oxide Synthase Is Not Essential for Control of Trypanosoma cruzi Infection in Mice

2004 ◽  
Vol 72 (7) ◽  
pp. 4081-4089 ◽  
Author(s):  
Kara L. Cummings ◽  
Rick L. Tarleton
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Trypanosoma Cruzi ◽  
Inducible Nitric Oxide Synthase ◽  
Interleukin 1 ◽  
Mouse Strains ◽  
Wild Type ◽  
Necrosis Factor Alpha ◽  
Oxide Synthase ◽  
Inducible Nitric Oxide

ABSTRACT Immune control of many intracellular pathogens, including Trypanosoma cruzi, is reported to be dependent on the production of nitric oxide. In this study, we show that mice deficient in inducible nitric oxide synthase (iNOS or NOS2) exhibit resistance to T. cruzi infection that is comparable to that of wild-type mice. This is the case for two iNOS-deficient mouse strains, Nos2tm1Lau and Nos2 N5, infected with the Brazil or Tulahuen strain of T. cruzi. In all cases, blood parasitemia, tissue parasite load, and survival rates are similar between wild-type and iNOS-deficient mice. In contrast, both wild-type and Nos2tm1Lau mice died within 32 days postinfection when treated with the nitric oxide synthase inhibitor aminoguanidine. Increased transcription of NOS1 or NOS3 is not found in iNOS-knockout (KO) mice, indicating that the absence of nitric oxide production through iNOS is not compensated for by increased production of other NOS isoforms. However, Nos2tm1Lau mice exhibit enhanced expression of tumor necrosis factor alpha, interleukin-1, and macrophage inflammatory protein 1α compared to that of wild-type mice, and these alterations may in part compensate for the lack of iNOS. These results clearly show that iNOS is not required for control of T. cruzi infection in mice.

NF-IL6 and AP-1 cooperatively modulate the activation of the TSG-6 gene by tumor necrosis factor alpha and interleukin-1

1994 ◽  
Vol 14 (10) ◽  
pp. 6561-6569
Author(s):  
L Klampfer ◽  
T H Lee ◽  
W Hsu ◽  
J Vilcek ◽  
S Chen-Kiang
Keyword(s):  
Tumor Necrosis Factor ◽  
Tumor Necrosis Factor Alpha ◽  
Tumor Necrosis ◽  
Interleukin 1 ◽  
Human Fibroblasts ◽  
Tnf Alpha ◽  
Necrosis Factor Alpha ◽  
Normal Human ◽  
Factor Alpha ◽  
Necrosis Factor

Tumor necrosis factor alpha (TNF-alpha) and interleukin-1 (IL-1) activate transcription of the TSG-6 gene in normal human fibroblasts through a promoter region (-165 to -58) that encompasses an AP-1 and a NF-IL6 site. We show by deletion analysis and substitution mutagenesis that both sites are necessary for activation by TNF-alpha. Activation by IL-1 requires the NF-IL6 site and is enhanced by the AP-1 site. These results suggest that the NF-IL6 and AP-1 family transcription factors functionally cooperate to mediate TNF-alpha and IL-1 signals. Consistent with this possibility, IL-1 and TNF-alpha markedly increase the binding of Fos and Jun to the AP-1 site, and NF-IL6 activates the native TSG-6 promoter. Activation by NF-IL6 requires an intact NF-IL6 site and is modulated by the ratio of activator to inhibitor NF-IL6 isoforms that are translated from different in-frame AUGs. However, the inhibitor isoform can also bind to the AP-1 site and repress AP-1 site-mediated transcription. The finding that the inhibitor isoform antagonizes activation of the native TSG-6 promoter by IL-1 and TNF-alpha suggests that NF-IL6 has a physiologic role in these cytokine responses. Thus, the functionally distinct NF-IL6 isoforms cooperate with Fos and Jun to positively and negatively regulate the native TSG-6 promoter by TNF-alpha and IL-1.

Induction of ICAM-1 by TNF-alpha, IL-1 beta, and LPS in human endothelial cells after downregulation of PKC

1992 ◽  
Vol 263 (4) ◽  
pp. C767-C772 ◽  
Author(s):  
C. L. Myers ◽  
S. J. Wertheimer ◽  
J. Schembri-King ◽  
T. Parks ◽  
R. W. Wallace
Keyword(s):  
Endothelial Cells ◽  
Protein Kinase ◽  
Umbilical Vein ◽  
Interleukin 1 ◽  
Protein Kinase Inhibitors ◽  
Intercellular Adhesion Molecule ◽  
Tnf Alpha ◽  
Necrosis Factor Alpha ◽  
Western Blots ◽  
Mrna Induction

The intercellular adhesion molecule 1 (ICAM-1) is induced on endothelial cells by tumor necrosis factor-alpha (TNF-alpha), interleukin-1 beta (IL-1 beta), and lipopolysaccharide (LPS). We have reported the sensitivity of cytokine-induced ICAM-1 expression to protein kinase inhibitors, including inhibitors of protein kinase C (PKC) [C. L. Myers, S. N. Desai, J. Schembri-King, G. L. Letts, and R. W. Wallace. Am. J. Physiol. 262 (Cell Physiol. 31): C365-C373, 1992]. To directly investigate the role of PKC in ICAM-1 induction, we downregulated PKC by pretreatment of human umbilical vein endothelial cells with phorbol 12-myristate 13-acetate (PMA) and assessed ICAM-1 protein and mRNA induction elicited by subsequent exposure to inflammatory stimuli. PMA treatment results in ICAM-1 protein induction that declines to basal levels by 3 days. Western blots of endothelial cell lysates reveal a nearly complete loss of immunologically reactive PKC. Subsequent activation with cytokine or LPS leads to reinduction of ICAM-1 protein and mRNA; however, the cells no longer produced substantial amounts of ICAM-1 protein or mRNA in response to PMA stimulation. Cross desensitization is observed with phorbol dibutyrate, while 4 alpha-phorbol has no desensitizing effect. The data indicate that PKC activation, while capable of inducing ICAM-1 expression, is not essential for ICAM-1 induction by the inflammatory mediators TNF-alpha, IL-1 beta, or LPS.

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