Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P43026 (lipopolysaccharide)
 62,215 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In heart muscle, the cytokine-inducible isoform of nitric oxide synthase (NOS2) is expressed in both cardiac myocytes and microvascular endothelial cells (CMEC). mRNA levels for both NOS2 and for osteopontin, a multifunctional extracellular matrix phosphoprotein containing and RGD integrin binding domain, are increased in cardiac muscle following intraperitoneal injection of adult rats with lipopolysaccharide. In vitro, interleukin-1 beta and interferon-gamma increased osteopontin mRNA levels in CMEC as well as NOS2 expression in both CMEC and cardiac myocytes. However, osteopontin mRNA levels in heart muscle in vivo, and in cardiac myocytes and CMEC in vitro, also are increased 10-30-fold by the synthetic glucocorticoid dexamethasone, an agent that suppresses cytokine induction of NOS2 in both cell types. The hexapeptide GRGDSP, which interrupts binding of RGD-containing proteins to cell surface integrins, increased NOS2 mRNA, while a synthetic osteopontin peptide analogue decreased NOS2 mRNA and protein levels in both cytokine-pretreated cardiac myocytes and CMEC cultures. Also, transfection with a full-length antisense-osteopontin cDNA in cytokine-pretreated CMEC decreased endogenous osteopontin mRNA and increased NOS2 mRNA levels. These results suggest that osteopontin could regulate the location and extent of NOS2 induction in the heart. Increased expression of osteopontin also may be one mechanism by which glucocorticoids suppress NOS2 activity in cardiac myocytes and microvascular endothelial cells.
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PMID:Glucocorticoids increase osteopontin expression in cardiac myocytes and microvascular endothelial cells. Role in regulation of inducible nitric oxide synthase. 749 54

Nitric oxide, a multifunctional effector molecule synthesized by nitric oxide synthase (NOS) from L-arginine, conveys signals for vasorelaxation, neurotransmission, and cytotoxicity. Three different NOS isoforms have been identified which fall into two distinct types, constitutive and inducible. The inducible NOS (iNOS) isoform is expressed in a variety of cell types and tissues in response to inflammatory agents and cytokines. The human iNOS (NOS2) gene was isolated on overlapping cosmid clones from a human genomic library using both the murine macrophage and the human hepatocyte iNOS cDNAs as probes. All isolated cosmids were part of a single genomic locus and no other genomic loci were identified or isolated. Analysis of this locus indicated that the human iNOS gene is approximately 37 kilobases in length and consists of 26 exons and 25 introns. Primer extension analysis of lipopolysaccharide and cytokine-stimulated human hepatocyte RNA mapped the transcriptional initiation site 30 base pairs downstream of a TATA sequence, and a 400-base pair 5'-flanking region was found to be structurally similar to the recently described murine iNOS promoter. Polymerase chain reaction analysis of a human/rodent genomic DNA somatic cell hybrid panel and fluorescent in situ hybridization indicated that the human iNOS gene is located on chromosome 17 at position 17cen-q11.2.
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PMID:Molecular cloning, structure, and chromosomal localization of the human inducible nitric oxide synthase gene. 750 10

The expression of inducible nitric oxide synthase (NOS2) is complex and is regulated in part by gene transcription. In this investigation we studied the regulation of NOS2 in a human liver epithelial cell line (AKN-1) which expresses high levels of NOS2 mRNA and protein in response to tumor necrosis factor alpha, interleukin 1 beta, and interferon gamma (cytokine mix, CM). Nuclear run-on analysis revealed that CM transcriptionally activated the human NOS2 gene. To delineate the cytokine-responsive regions of the human NOS2 promoter, we stimulated AKN-1 cells with CM following transfection of NOS2 luciferase constructs. Analysis of the first 3.8 kb upstream of the NOS2 gene demonstrated basal promoter activity but failed to show any cytokine-inducible activity. However, 3- to 5-fold inductions of luciferase activity were seen in constructs extending up to -5.8 and -7.0 kg, and a 10-fold increase was seen upon transfection of a -16 kb construct. Further analysis of various NOS2 luciferase constructs ligated upstream of the thymidine kinase promoter identified three regions containing cytokine-responsive elements in the human NOS2 gene: -3.8 to -5.8, -5.8 to -7.0, and -7.0 to -16 kb. These results are in marked contrast with the murine macrophage NOS2 promoter in which only 1 kb of the proximal 5' flanking region is necessary to confer inducibility to lipopolysaccharide and interferon gamma. These data demonstrate that the human NOS2 gene is transcriptionally regulated by cytokines and identify multiple cytokine-responsive regions in the 5' flanking region of the human NOS2 gene.
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PMID:Transcriptional regulation of human inducible nitric oxide synthase (NOS2) gene by cytokines: initial analysis of the human NOS2 promoter. 857 13

Induction of hepatic nitric oxide synthase (NOS) by tumor necrosis factor-alpha (TNF alpha), interleukin-1 beta (IL-1 beta), interferon-gamma (IFN gamma), interleukin-6 (IL-6), and lipopolysaccharide was assessed as activity and immunoreactive protein. Hepatic NOS activity was cytosolic and had cofactor requirements consistent with inducible nitric oxide synthase (NOS2). NOS induction by TNF alpha was dose dependent from concentrations of 0.06 to 60 nM and was increased 2-3-fold by IFN gamma. NOS induction was reflective of total TNF alpha binding to hepatocyte receptors. Hepatocyte TNF alpha binding fit a biphasic curve with high affinity (K(d) = 1.4 nM, Bmax = 3157 sites) and low affinity (K(d) = 157 nM, Bmax = 204,948 sites) elements. NOS2 activity was induced by lipopolysaccharide, IL-1 beta, TNF alpha, and IFN gamma but not by IL-6. All cytokine stimuli were inhibited by antioxidants. Oxygen radical generation was directly measured as dichlorofluoroscein fluorescence in isolated mitochondria. Mitochondria from TNF alpha-treated hepatocytes generated more oxygen radicals than did controls. Antioxidants reduced mitochondrial generation of oxygen radicals. Activation of the transcription factor nuclear factor-kappa B by TNF alpha, IFN gamma, and IL-1 beta was assessed by gel shift analysis. Cytokine treatment increased nuclear factor-kappa B binding, and the addition of antioxidants or rotenone inhibited cytokine activation. Taken together, these data suggest that oxygen radicals, possibly generated by mitochondria, play a major role in NOS2 induction by cytokines.
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PMID:Characterization of hepatic nitric oxide synthase: identification as the cytokine-inducible form primarily regulated by oxidants. 870 Jan 34

The roles of nitric oxide derived from either the constitutive endothelial NO synthase (eNOS or NOS3) or the inducible NOS (iNOS or NOS2) in hepatic injury during endotoxemia remain controversial. To investigate this further, rats received a bolus of lipopolysaccharide (LPS) following implantation of osmotic pumps containing one of two nonselective NOS inhibitors (NMA or NAME), one of two inducible NOS inhibitors (NIL or AG), or saline. The inhibitors were infused continuously into the liver via the portal vein. Treatment of LPS-injected rats with NMA and NAME resulted in 106 and 227% increases, respectively, in circulating hepatic enzyme levels compared to LPS-treated control rats. In contrast, infusion of the iNOS-selective inhibitors had no effect on the LPS-induced hepatic necrosis. In rats receiving NAME, LPS induced greater neutrophil infiltration and ICAM-1 expression than in the LPS + saline group, whereas NIL infusion did not. The increased hepatic necrosis and PMN infiltration in the LPS + NAME group was partially prevented by a simultaneous infusion of a liver-selective NO donor. Inhibition of PMN accumulation using an anti-ICAM-1 antibody or by PMN depletion using vinblastine pretreatment, however, did not reverse the increased necrosis with NAME infusion during endotoxemia. In contrast to the assessment for necrosis, increased apoptosis was observed in the livers of LPS-treated rats receiving infusions of either NAME or NIL, but not with LPS alone. These data indicate that NO produced by eNOS may be adequate to prevent necrosis by a mechanism independent of PMN, while induced NO appears to prevent apoptosis.
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PMID:Differential effects of nonselective nitric oxide synthase (NOS) and selective inducible NOS inhibition on hepatic necrosis, apoptosis, ICAM-1 expression, and neutrophil accumulation during endotoxemia. 944 11

In previous studies we reported that stimulation of rat mesangial cells (RMC) with lipopolysaccharide (LPS) + tumor necrosis factor alpha (TNF-alpha) (L/T) elicits inducible nitric oxide synthase (NOS2) mRNA expression, which is inhibited by dexamethasone (DX). We have now analyzed the mechanisms responsible for this inhibitory effect. Dexamethasone had no destabilizing effect on NOS2 mRNA. Transfection of RMC with several luciferase reporter constructs from the 5' flanking regulatory region of the rat NOS2 gene established the importance of the NF-kappa B site in the transcriptional activation of the NOS2 gene. DNA mobility shift assays showed activation by L/T of the NF-kappa B complex in a time-dependent manner. Dexamethasone specifically inhibited this activation in a process dependent on the glucocorticoid receptor and with a markedly greater effect when it was added prior to L/T. Dexamethasone increased the expression of the I kappa B-alpha transcript and protein in the cytoplasm. While treatment of RMC with L/T induced the transient decrement of cytoplasmic p65 levels and its appearance in the nucleus, preincubation with DX prevented this effect. Co-immunoprecipitation and immunocytochemical studies demonstrated that I kappa B-alpha is associated with p65 in the cytoplasm of RMC after treatment with DX and L/T. These results prove that inhibition of NF-kappa B-mediated transcription is a crucial mechanism by which DX inhibits NOS2 expression, and that this occurs by increasing cytoplasmic I kappa B-alpha levels and sequestering the activating subunits of NF-kappa B in the cytoplasm. The need for previous induction of I kappa B-alpha could provide a molecular explanation for the limited efficacy of these agents in the therapy of septic shock.
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PMID:Involvement of transcriptional mechanisms in the inhibition of NOS2 expression by dexamethasone in rat mesangial cells. 945 98

The expression of the inducible isoform of nitric oxide synthase (NOS2, iNOS) is increased in patients undergoing sepsis as well as in animal models in which septic shock is induced by injection of bacterial lipopolysaccharide (LPS). Transforming growth factor-beta1 (TGF-beta1) potently suppresses NO production both in vitro and in vivo. After intraperitoneal injection of LPS, mice over-expressing a cDNA coding for active TGF-beta1 in the liver (Alb/ TGF-beta1) exhibited reduced serum levels of the NO reaction products NO2(-) + NO3(-) compared with controls. Paradoxically, while endotoxemic Alb/ TGF-beta1 mice expressed much less NOS2 protein in peritoneal exudate cells than did endotoxemic wild-type mice, Alb/TGF-beta1 mice expressed more NOS2 mRNA and protein in both liver and kidney. Alb/ TGF-beta1 mice treated with LPS had eightfold higher serum tumor necrosis factor alpha (TNF-alpha) levels and experienced increased mortality compared with wild-type mice, which was associated with renal insufficiency. These results suggest that renal dysfunction, decreased production of NO, and/or increased production of TNF-alpha are associated with increased mortality of endotoxemic Alb/TGF-beta1 mice.
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PMID:Increased mortality, blunted production of nitric oxide, and increased production of TNF-alpha in endotoxemic TGF-beta1 transgenic mice. 946 70

Macrophages are activated to become cytotoxic by a highly coordinated set of cytokine signals. Ionizing radiation can mimic cytokine signals and lead to enhanced states of activation. We tested the ability of gamma-radiation, alone and with interferon-gamma (IFN-gamma) and/or lipopolysaccharide (LPS), to induce nitric oxide (NO) production in J774.1 and RAW264.7 murine macrophages. NO was induced weakly, moderately, or strongly by IFN-gamma alone, LPS alone, or IFN-gamma + LPS, respectively. Radiation alone (0.5-50 Gy) did not induce NO, but enhanced NO production in a dose-dependent manner (0.5-5 Gy) when cells were exposed to IFN-gamma or LPS 24 h post-irradiation. Immunoblots showed parallel induction of nitric oxide synthase (NOS2). Application of anti-tumor necrosis factor alpha (TNF-alpha) antibody before irradiation blocked induction of NO by IFN-gamma. We conclude (1) that irradiated cells produce more NO in response to either IFN-gamma or LPS and (2) that the increase is mediated by induction of TNF-alpha.
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PMID:Ionizing radiation potentiates the induction of nitric oxide synthase by IFN-gamma and/or LPS in murine macrophage cell lines: role of TNF-alpha. 976 26

Inducible synthesis of nitric oxide (NO) by macrophages is an important mechanism of the host defense against intracellular infection in mice, but the evidence for significant levels of inducible NO production by human macrophages is controversial. Here we report that the human promyelocytic cell line HL-60, when differentiated to a macrophage-like phenotype, acquires the ability to produce substantial amounts of NO on stimulation with LPS or 1, 25-dihydroxyvitamin D3 (1,25-D3) in the absence of activating factors such as gamma interferon. Expression of the inducible nitric oxide synthase (NOS2) was confirmed by sequencing of the reverse transcription-PCR product from stimulated HL-60 cells. Kinetic studies after lipopolysaccharide stimulation show that NOS2 mRNA levels rise within 3 to 6 h, that conversion of [14C]arginine to [14C]citrulline is maximal at 5 to 6 days, and that levels of reactive nitrogen intermediates stabilize at around 20 microM at 7 to 8 days. We find that 1,25-D3 acts to suppress the growth of Mycobacterium tuberculosis in these cells and that this effect is inhibited by NG-monomethyl-L-arginine, suggesting that vitamin D-induced NO production may play a role in the host defense against human tuberculosis.
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PMID:1,25-Dihydroxyvitamin D3 induces nitric oxide synthase and suppresses growth of Mycobacterium tuberculosis in a human macrophage-like cell line. 978 38

Nitric oxide (NO) production in macrophages by inducible nitric oxide synthase (NOS2) has multiple tissue damaging effects and is involved in the pathogenesis of inflammation and graft rejection. Haem oxygenase (HmOx) is the enzyme which degrades haem. Its inducible isoform, HmOx1, was recently shown to increase cellular resistance against oxidative stress and to decrease inflammation and graft rejection. Since haem is an essential cofactor for NOS2 activity, we investigated the effects of HmOx1-induction upon NO secretion in macrophages. We induced HmOx1 in BALB/c bone-marrow-derived macrophages by short-term exposure to haemin (20 micromol/l, 30 min); then we incubated them for 24 h to allow maximal expression of HmOx1 activity. Next, we activated the macrophages with lipopolysaccharide (LPS) and measured their NO production and their NO-dependent cytotoxicity against P815 cells. We found that HmOx induction 24 h before LPS activation in mouse macrophages suppresses their production of NO, while HmOx inhibition (with zinc protoporphyrin) increases NO secretion. NOS2 inhibition is reflected by the decrease of macrophage NO-dependent cytotoxicity against the P815 targets. We therefore propose that HmOx1 is a physiological inhibitor of NOS2 in activated macrophages because it decreases haem availability for NOS2 synthesis. NOS2 inhibition may explain the antinflammatory effects of HmOx induction which could also be used therapeutically in situations when NO hyperproduction leads to cytotoxic effects such as inflammation or transplant rejection.
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PMID:Nitric oxide synthase inhibition by haem oxygenase decreases macrophage nitric-oxide-dependent cytotoxicity: a negative feedback mechanism for the regulation of nitric oxide production. 985 35


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