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)

Three isozymes of nitric oxide (NO) synthase (EC 1.14.13.39) have been identified and the cDNAs for these enzymes isolated. In humans, isozymes I (in neuronal and epithelial cells), II (in cytokine-induced cells), and III (in endothelial cells) are encoded for by three different genes located on chromosomes 12, 17, and 7, respectively. The deduced amino acid sequences of the human isozymes show less than 59% identity. Across species, amino acid sequences for each isoform are well conserved (> 90% for isoforms I and III, > 80% for isoform II). All isoforms use L-arginine and molecular oxygen as substrates and require the cofactors NADPH, 6(R)-5,6,7,8-tetrahydrobiopterin, flavin adenine dinucleotide, and flavin mononucleotide. They all bind calmodulin and contain heme. Isoform I is constitutively present in central and peripheral neuronal cells and certain epithelial cells. Its activity is regulated by Ca2+ and calmodulin. Its functions include long-term regulation of synaptic transmission in the central nervous system, central regulation of blood pressure, smooth muscle relaxation, and vasodilation via peripheral nitrergic nerves. It has also been implicated in neuronal death in cerebrovascular stroke. Expression of isoform II of NO synthase can be induced with lipopolysaccharide and cytokines in a multitude of different cells. Based on sequencing data there is no evidence for more than one inducible isozyme at this time. NO synthase II is not regulated by Ca2+; it produces large amounts of NO that has cytostatic effects on parasitic target cells by inhibiting iron-containing enzymes and causing DNA fragmentation. Induced NO synthase II is involved in the pathophysiology of autoimmune diseases and septic shock. Isoform III of NO synthase has been found mostly in endothelial cells. It is constitutively expressed, but expression can be enhanced, eg, by shear stress. Its activity is regulated by Ca2+ and calmodulin. NO from endothelial cells keeps blood vessels dilated, prevents the adhesion of platelets and white cells, and probably inhibits vascular smooth muscle proliferation.
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PMID:Nitric oxide synthase isozymes. Characterization, purification, molecular cloning, and functions. 751 53

Nitric oxide (NO) produced by endothelial cells (EC) has been shown to exert cytotoxic activity on tumor cells. In order to analyze events involved in brain metastasis, the modulation of NO production in rat-brain-derived EC was investigated. NO release was increased by tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma), interleukin-1 beta, lipopolysaccharide or forskolin in EC219 cells, a rat-brain-microvessel-derived EC line. Dexamethasone decreased NO release by cytokine-activated EC219 cells. Tumor cells (DHD/K12/PROb, a rat colon-carcinoma cell line) were highly adherent to EC219 cells, and adhesion was not modified by TNF-alpha plus IFN-gamma, or by dexamethasone. Addition of tumor cells or tumor-cell-conditioned medium significantly inhibited NO release induced by any of the stimuli examined, but only if added during the initial phase of endothelial-cell activation. Tumor-derived suppression of NO release was also observed in primary cultures of cerebral EC. NO synthase (NOS) activity in cytosol extracts of the cerebral EC line was Ca(2+)-independent and required both NADPH and tetrahydrobiopterin. NOS activity was increased by TNF-alpha and IFN-gamma, and significantly reduced by tumor-cell-conditioned medium. These results suggest that rat colon-carcinoma cells may have developed a protective mechanism involving the release of (a) soluble factor(s) which inhibit(s) NO production by cerebral EC.
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PMID:Tumor cells suppress cytokine-induced nitric-oxide (NO) production in cerebral endothelial cells. 752 97

Nitric oxide (NO) synthase activity was detected in fat body and the Malpighian tubles of the silkworm, Bombyx mori. Main NO synthase activity in the fat body was Ca(2+)/calmodulin-dependent, inducible by bacterial lipopolysaccharide (LPS) and required NADPH, FAD, FMN, dithiothreitol (DTT) and tetrahydrobiopterin (BH4) as cofactors for the full expression of the activity. The Malpighian tubles contained two types of NO synthase. One was Ca(2+)-independent, calmodulin-dependent and constitutive and the other was Ca(2+)-dependent and constitutive. The former NO synthase required the same cofactors as fat body NO synthase. The activity of Malpighian tuble NO synthases increased dramatically at the end of the last instar period, just prior to spinning. These results indicate that B. mori contains new types of NO synthase, suggesting the wide distribution and different characteristics of this enzyme among vertebrates and invertebrates.
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PMID:Occurrence of novel types of nitric oxide synthase in the silkworm, Bombyx mori. 753 73

Nitric oxide (NO) produces rapid osteoclast detachment and contraction in vitro, and this effect is accompanied by a profound inhibition of bone resorption. Work by others has confirmed these findings in vivo: inhibition of NO synthase [NOS; L-arginine, NADPH: oxygen oxidoreductase (NO-forming), EC 1.14.13.39] in normal rats is followed by increased bone resorption reflected by a marked loss in bone mineral density. In our present study, immunocytochemistry and Northern blotting show the presence of the constitutive calcium-sensitive NOS isoform (cNOS) in normal rat osteoclasts and in the human preosteoclast cell line (FLG 29.1). The inducible NOS isoform (iNOS) was also clearly demonstrable in the rat cells especially after treatment with gamma interferon (IFN-gamma) and bacterial wall products [lipopolysaccharide (LPS)], while a basal level of transcript was detected in the untreated human preosteoclast line. However NADPH-diaphorase activity was intense only in neonatal rat osteoclasts attached to bone, perhaps reflecting either enhancement of cNOS activity by calcium or increased amounts of the inducible isoform in activated osteoclasts in situ compared with isolated neonatal rat osteoclasts. These actively resorb devitalized bone but the untreated cells contain relatively low levels of NOS; they are extremely sensitive to inhibition by NO. The iNOS inhibitor aminoguanidine markedly enhances in vitro resorption by activated NOS-rich chick osteoclasts and by normal rat osteoclasts treated with LPS or IFN-gamma. In contrast, the nonselective NOS inhibitor NG-monomethyl-L-arginine inhibits resorption by untreated neonatal rat osteoclasts. Thus, osteoclast function may require intermittent calcium-stimulated increases in NO production by cNOS against a basal inhibitory background activity of the iNOS isoform. However, bone resorption depends on precursor replication and on the activity of the mature cells, and we found that the NO donor 3-morpholinosydnonimine (SIN-1) (50 microM) profoundly depressed replication in the human preosteoclast line. Taken together, these results strongly suggest that NO maintains a central control of bone resorption in both avian and mammalian species by exerting a powerful tonic restraint of osteoclast numbers and activity. The presence of NOS in human cells implies a similar function in man and that conventional views of calcium homoeostasis and skeletal metabolism will need substantial revision. Since NO also influences behavior of the osteoblast, the bone-forming cell, in vitro, a similar effect in vivo might imply a general influence on bone remodeling.
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PMID:Bidirectional regulation of osteoclast function by nitric oxide synthase isoforms. 753 33

The prevailing view of neutrophil NADPH-oxidase activation during interaction with bacteria is that the production of toxic oxygen metabolites should be directed into the phagosome containing the engulfed prey. However, in this report we show that a common Escherichia coli strain, HB101, may induce a release of neutrophil oxygen metabolites to the extracellular milieu. This phenomenon is dependent on three factors: (i) the mobilization (upregulation) of neutrophil secretory vesicles prior to interaction with the bacteria, (ii) soluble bacterial factors binding to the formylmethionyl-leucyl-phenylalanine receptor and tentatively identified as formylated peptides, and (iii) a bacterium-associated priming factor identified as lipopolysaccharide.
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PMID:Escherichia coli-induced activation of neutrophil NADPH-oxidase: lipopolysaccharide and formylated peptides act synergistically to induce release of reactive oxygen metabolites. 759 Nov 13

This study was performed to determine the magnitude and time of onset of in vivo changes in hepatic bioenergetics in response to a sublethal dose of lipopolysaccharide (LPS), a bacterial endotoxin. Male rats (48-hour-fasted) were administered an intraperitoneal injection of LPS (5 mg/kg body weight) or vehicle alone, and the livers were freeze-clamped 5, 30, or 180 minutes or 24 hours later. Liver tissue was extracted with perchloric acid, and the metabolites necessary to calculate NAD(+)- and NADP(+)-linked redox states and the cytosolic phosphorylation potential were measured. There was no significant difference in hepatic cytosolic phosphorylation potential between LPS and control groups at any of the times investigated. This indicated that the ability of the liver to synthesize adenosine triphosphate (ATP) was not compromised under the conditions of the study. No changes in hepatic redox states were observed 5 or 30 minutes after LPS treatment. Three hours after LPS treatment, hepatic cytosolic and mitochondrial free-[NAD+]/[NADH] redox states and the cytosolic free-[NADP+]/[NADPH] redox state were more oxidized. By 24 hours, only NAD(+)-linked redox states were more oxidized than the time-matched controls. Hepatic urea content was elevated at both 3 and 24 hours, compatible with an increased rate of urea synthesis as a consequence of increased amino acid metabolism, whereas hepatic beta-hydroxybutyrate and total ketone bodies were decreased 24 hours after LPS treatment, indicating decreased hepatic ketogenesis.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:In vivo effects of lipopolysaccharide on hepatic free-NAD(P)(+)-linked redox states and cytosolic phosphorylation potential in 48-hour-fasted rats. 766 91

Stimulation of nitric oxide (NO) synthase in endothelial cells by Ca2+ influx leads to increased intracellular levels of cGMP. NO synthase from various sources is known to use tetrahydrobiopterin, flavins, and NADPH as cofactors. We studied the effect of interferon-gamma, tumor necrosis factor-alpha, and lipopolysaccharide on tetrahydrobiopterin biosynthetic activities in human umbilical vein endothelial cells (HUVEC). These stimuli led to an up to 40-fold increase of GTP cyclohydrolase I (EC 3.5.4.16) activity and to increased accumulation of neopterin and tetrahydrobiopterin in HUVEC. Further enzyme activities of tetrahydrobiopterin biosynthesis, i.e. 6-pyruvoyl tetrahydropterin synthase and sepiapterin reductase (EC 1.1.1.153), remained unchanged. NO synthase activity in protein fractions from homogenates of cells treated with interferon-gamma plus tumor necrosis factor-alpha was not influenced as compared with untreated controls. However, interferon-gamma alone or in combination with tumor necrosis factor-alpha significantly increased intracellular cGMP formation in intact HUVEC by 50 and 80%, respectively. These stimuli increased intracellular tetrahydrobiopterin concentrations up to 14-fold. NO-triggered cGMP formation was similarly increased by incubation of otherwise untreated cells with sepiapterin, leading to elevated intracellular tetrahydrobiopterin levels. Thus, cytokines indirectly stimulate the activity of constitutive NO synthase in HUVEC by upregulating production of the cofactor tetrahydrobiopterin.
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PMID:Pteridine biosynthesis in human endothelial cells. Impact on nitric oxide-mediated formation of cyclic GMP. 767 11

Nitric oxide is a short-lived biologic mediator for diverse cell types. Synthesis of an inducible nitric oxide synthase (NOS) in murine macrophages is stimulated by lipopolysaccharide (LPS) and interferon gamma. In human hepatocytes, NOS activity is induced by treatment with a combination of tumor necrosis factor, interleukin 1, interferon gamma, and LPS. We now report the molecular cloning and expression of an inducible human hepatocyte NOS (hep-NOS) cDNA. hep-NOS has 80% amino acid sequence homology to macrophage NOS (mac-NOS). Like other NOS isoforms, recognition sites for FMN, FAD, and NADPH are present, as well as a consensus calmodulin binding site. NOS activity in human 293 kidney cells transfected with hep-NOS cDNA is diminished by Ca2+ chelation and a calmodulin antagonist, reflecting a Ca2+ dependence not evident for mac-NOS. Northern blot analysis with hep-NOS cDNA reveals a 4.5-kb mRNA in both human hepatocytes and aortic smooth muscle cells following stimulation with LPS and cytokines. Human genomic Southern blots probed with human hep-NOS and human endothelial NOS cDNA clones display different genomic restriction enzyme fragments, suggesting distinct gene products for these NOS isoforms. hep-NOS appears to be an inducible form of NOS that is distinct from mac-NOS as well as brain and endothelial NOS isozymes.
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PMID:Molecular cloning and expression of inducible nitric oxide synthase from human hepatocytes. 768 6

1. Male Sprague-Dawley or Wistar rats were injected with bacterial lipopolysaccharide (LPS; 5 mg kg-1, i.p.) and killed after 1, 3, 6, 15, and 24 h. The brains, mesenteries, spleens, lungs, livers, kidneys, hearts, aortae and diaphragms were removed and frozen immediately. Control rats were injected with sterile saline and killed after 6 h. 2. The organs were homogenized in a semi-frozen state and NO synthase (NOS) activity measured in tissues from both LPS-treated and saline-treated groups by the ability of homogenates to convert [3H]-L-arginine to [3H]-L-citrulline in a NADPH-dependent manner. 3. The NOS activity in all organs taken from control animals was found to be calcium-dependent, with the highest activity being in the brain. After LPS-treatment an induced calcium-independent NOS was detected in all tissues tested, with the exception of the brain. The spleen, lung, mesentery and liver had the highest amounts of LPS-induced NOS activity. No induction of calcium-dependent NOS was detected. 4. Induction of NOS was maximum 6 h after administration of LPS and had returned to control levels in 24 h. 5. The constitutive NOS in brain and mesentery and the LPS-induced activities in the spleen, lung, liver and mesentery were inhibited by NG-monomethyl-L-arginine (L-NMMA) or NG-nitro-L-arginine methyl ester (L-NAME) according to concentration. The IC50 for L-NAME was 2.5 microM against the constitutive NOS from brain, and 20-25 microM against the inducible NOS. For L-NMMA the IC50 was 20-25 microM against either NOS isoform. 7. The vascular responses to endothelin-I (ET-1), the thromboxane A2-mimetic 11 alpha,9 alpha-epoxymethanoprostaglandin F2alpha (U46619), phenylephrine (PE) or 5-hydroxytryptamine (5-HT) were measured in the simultaneously perfused arterial and venous mesenteric vascular beds from both control and LPS-treated(6 h) rats. Vasoconstrictor responses to all agonists tested were unaffected by LPS treatment. In the presence of L-NAME (100 microM) vasoconstrictor responses were potentiated in both the arterial and venous portion of the mesenteric beds from both control and LPS-treated rats. The potentiation of responses to U46619 was significantly greater in beds from LPS-treated rats.8. Injection of LPS i.p. is associated with induction of NOS in all organs tested, except for the brain. In the mesentery this is not accompanied by a hyporesponsiveness to constrictor agents suggesting an increased sensitivity, particularly to U46619. This may explain the poor perfusion and tissue damage in the splanchnic circulation associated with sepsis.
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PMID:Induction by endotoxin of nitric oxide synthase in the rat mesentery: lack of effect on action of vasoconstrictors. 768 6

Calmodulin-dependent nitric-oxide synthase, with an apparent molecular mass of 125 kDa, was induced in the liver of rats treated with Propionibacterium acnes and Escherichia coli lipopolysaccharide. Clones were isolated from a cDNA library obtained from induced rat liver using oligonucleotide probes which were synthesized based on the amino acid sequences of peptides of the purified enzyme. Four overlapping cDNA clones for a 3.8-kbp region were isolated and the nucleotide sequences were determined. These clones encompassed an open-reading frame of 3441 bases encoding 1147 amino acids. The deduced amino acid sequence of the cDNA suggested that the protein contains binding sites for NADPH, FAD and FMN. The structure of the possible calmodulin-binding site, consisting of a strongly hydrophobic region surrounded by basic amino acids, is present. The full-length cDNA was expressed in COS 1 cells under the control of a cytomegalovirus promoter and the expressed enzyme was found to be a calmodulin-dependent nitric-oxide synthase. A structural comparison suggested that the liver nitric-oxide synthase is the same as the macrophage enzyme. Northern-blot analysis showed that the mRNA in the liver is approximately 4.2 kb long and is induced transcriptionally by treatment with P. acnes and lipopolysaccharide.
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PMID:Molecular cloning of a cDNA encoding an inducible calmodulin-dependent nitric-oxide synthase from rat liver and its expression in COS 1 cells. 769 62


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