Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

The principal goal of the present study was to test the hypothesis that cytokines modulate glucose transport in skeletal muscle by increasing nitric oxide production. Cultured L6 skeletal muscle cells were incubated in the presence of tumour necrosis factor-alpha, interferon-gamma or lipopolysaccharide (LPS) alone or in combination for 24 h. Neither cytokines nor LPS alone induced NO production, as measured by nitrite concentrations in the medium. However, when used in combination, the two cytokines significantly stimulated NO production, and this effect was synergistically enhanced by the presence of LPS. Reverse transcriptase-PCR (RT-PCR) analysis revealed that NO release was associated with the induction of inducible (macrophage-type) NO synthase (iNOS). The increase in iNOS expression was confirmed at the protein level by Western-blot analysis and NADPH/diaphorase histochemical staining. Cytokines and LPS markedly increased basal glucose transport in L6 myocytes. Insulin also stimulated basal glucose transport, but significantly less in cells chronically exposed to cytokines/LPS. The sensitivity of L6 muscle cells to insulin-stimulated glucose transport was also significantly decreased by cytokines/LPS treatment. The NOS inhibitor NG-nitro-l-arginine methyl ester (l-NAME) inhibited nitrite production in cytokine/LPS-treated cells, and this prevented the increase in basal glucose transport and restored muscle cell responsiveness to insulin. Cytokines/LPS exposure significantly increased GLUT1 transporter protein levels but decreased GLUT4 expression in L6 cells. l-NAME treatment prevented the increase in GLUT1 protein content but failed to restore GLUT4 transporter levels. These results demonstrate that cytokines and LPS affect glucose transport and insulin action by inducing iNOS expression and NO production in skeletal muscle cells. The data further indicate that cytokines and LPS increase the expression of the GLUT1 transporter protein by an NO-dependent mechanism.
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PMID:Cytokines modulate glucose transport in skeletal muscle by inducing the expression of inducible nitric oxide synthase. 923 Jan 32

Nitric oxide synthase (NOS) is responsible for the biological production of nitric oxide (NO) in several organs. NOS activity has also been localized in the reproductive tract, although direct evidence for its presence in the human or bovine oviduct is still lacking. In the present study, four different techniques were used to identify the presence of NOS activity in human (n = 11) and bovine (n = 9) oviduct: (i) conversion of [3H]-L-arginine to [3H]-L-citrulline; (ii) production of nitrite/nitrate (NO2/NO3; stable NO metabolites); (iii) identification of NADPH-diaphorase activity; and (iv) immunostaining with antiserum to endothelial NOS. Cytosolic extracts from human ampullary segments of the Fallopian tube, obtained from post-partum patients (n = 4), converted [3H]-L-arginine to [3H]-L-citrulline (21.0 +/- 8.8 fmol/mg protein/min). This conversion rate was significantly (P < 0.05) reduced in the presence of either EDTA or N-monomethyl-L-arginine monoacetate (L-NMMA), an inhibitor of NOS activity. When bovine (n = 3) ampullary segments were incubated for 36 h in Hanks' balanced salt solution, the concentration of NO2/NO3 in the medium was increased (P < 0.05) if segments were pretreated with lipopolysaccharide (LPS; an inducer of inducible NOS), but not after treatment with LPS + L-NMMA. Additionally, epithelial cells cultured from ampullary segments showed positive staining both for NADPH-diaphorase activity and with antiserum to endothelial NOS. The results of the present study provide direct evidence for the presence of both the Ca(2+)-dependent constitutive form of NOS, as well as the inducible form of NOS activity in human and bovine oviduct. Since the oviduct plays a key role in the reproductive process, it is possible that the two forms of NOS may be involved in the physiological regulation of oviduct function.
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PMID:Identification of nitric oxide synthase in human and bovine oviduct. 923 73

Synthesis of nitric oxide (NO) has been shown in the glandular epithelium of human prostate, with highest levels in the peripheral zone. This location is believed to be the main source of prostatic cancer. The ability of stromal cells to produce NO may contribute to the malignant process. Since solid tumours are prone to hypoxia and malignant progression, experiments were undertaken to test the effect of respiratory block on the induction of nitric oxide synthase (NOS) by a Dunning rat prostatic epithelial line. A metastatic phenotype (Mat-LyLu) was treated in vitro with brief exposure to cyanide in order to mimic transient hypoxic stress. NADPH-diaphorase activities in paraformaldehyde-fixed cells was used to follow the expression of NOS. NADPH-diaphorase activity was found to be inducible by a range of factors, including mechanical damage and infection of cultures. Cyanide induced a dose-dependent staining that was statistically greater than in untreated cells. Consistent with diaphorase staining being a marker for the inducible isoform of NOS (iNOS), induction and enhancement of staining, respectively, was observed in response to treatment with lipopolysaccharide or withdrawal of dexamethasone supplement. Results demonstrate that prostatic epithelia can be triggered in culture to express iNOS by transient oxidative stress in the form of respiratory poisoning by NaCN. Paradoxically, nitric oxide production by epithelia within hypoxic zones of solid tumours may contribute to the promotion and/or inhibition of tumorigenesis.
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PMID:Transient block of respiratory chain by cyanide triggers NADPH-diaphorase activity (a marker for nitric oxide synthase) in Dunning rat prostatic epithelium. 945 79

Inducible nitric oxide (NO) synthase (iNOS)-mediated hyperproduction of NO in airways has been reported in asthmatic patients. However, the role of NO in the pathogenesis of asthma has not yet been fully elucidated. The aim of this study was to examine whether the iNOS-derived NO affects airway microvascular leakage, one of the characteristic features of asthmatic airway inflammation. Guinea-pigs were exposed to lipopolysaccharide (LPS) (1 mg x mL(-1)) by inhalation in order to induce iNOS in the airways, and the histochemical staining of reduced nicotinamide-adenine dinucleotide phosphate (NADPH)-diaphorase activity was determined 5 h after the inhalation to confirm the iNOS induction. Airway microvascular leakage to subthreshold doses of substance P (0.3 microg x kg(-1), i.v.) was also examined in the absence and presence of an iNOS inhibitor (aminoguanidine) in LPS- or saline-exposed (control) animals using Evans blue dye and Monastral blue dye. In the LPS-exposed animals, increased NADPH-diaphorase activity was observed in the airway microvasculature compared with the control animals. Substance P caused significant airway microvascular leakage assessed by Evans blue dye in all airway levels in the LPS-exposed animals but not in the control group. This was also confirmed by Monastral blue dye extravasation. Aminoguanidine abolished this LPS-induced enhancement of plasma leakage to substance P without changing the systemic blood pressure. These results may suggest that inducible nitric oxide synthase-derived nitric oxide is capable of potentiating neurogenic plasma leakage in airways.
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PMID:Induction of nitric oxide synthase by lipopolysaccharide inhalation enhances substance P-induced microvascular leakage in guinea-pigs. 981 54

We investigated the pathophysiological role of nitric oxide synthesized by inducible nitric oxide synthase in the brain, by injecting lipopolysaccharide directly into the rat cerebral cortex/hippocampus. The levels of nitric oxide metabolites, nitrite and nitrate, began to increase in a dose-dependent manner with a 3-h lag, and reached approximately seven-fold the basal levels 8 h after the direct injection of lipopolysaccharide (5 microg). The lipopolysaccharide-induced increase in nitrite and nitrate levels was inhibited by treatment with the specific inducible nitric oxide synthase inhibitor aminoguanidine. The protein synthesis inhibitor cycloheximide delayed the onset of the increase in nitric oxide metabolite levels, and reduced the peak levels. Lipopolysaccharide increased Ca2+-independent, but not Ca2+-dependent, nitric oxide synthase activity in the brain. Intense nicotinamide adenine dinucleotide phosphate-diaphorase activity was observed in round cells in the vicinity of the site of injection of lipopolysaccharide 8 h after the injection. Neuronal death was observed seven days after the injection of lipopolysaccharide. Spatial memory, as assessed by performance in a water maze task and spontaneous alternation behavior in a Y-maze, was significantly impaired in rats which had had previous bilateral injections of lipopolysaccharide into the hippocampus. The lipopolysaccharide-induced neuronal death and spatial memory impairments were prevented by aminoguanidine. These results suggest that direct injection of lipopolysaccharide into the brain causes an induction of inducible nitric oxide synthase in vivo. Furthermore, it is suggested that nitric oxide produced by inducible nitric oxide synthase is responsible for the lipopolysaccharide-induced brain dysfunction.
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PMID:Brain dysfunction associated with an induction of nitric oxide synthase following an intracerebral injection of lipopolysaccharide in rats. 1005 Dec 7

Nitric oxide (NO) is known to be involved in the modulation of neuroendocrine function. To clarify the role of different isoforms of NO synthase (NOS) in the neuroendocrine response to immune challenge, the expressions of neuronal NOS (nNOS) and inducible NOS (iNOS) genes in the hypothalamus following lipopolysaccharide (LPS) injection were examined using in situ hybridization. NOS activity was also determined by NADPH-diaphorase (NADPH-d) histochemistry. LPS (25 mg/kg) or sterile saline was injected intraperitoneally to male Wistar rats and the rats sacrificed 30 min, or 1, 2, 3, 5, 12 or 24 h after injection. nNOS mRNA expression in the paraventricular nucleus (PVN) was significantly increased 2 h after LPS injection. iNOS mRNA, which was not detected until 2 h after LPS injection, was significantly increased in the PVN 3 h after LPS injection. Both RNA expressions had returned to basal levels by 12 h after LPS injection. The number of NADPH-d positive cells was significantly increased 5 h after LPS injection. iNOS expression was more robust in parvocellular PVN, while nNOS was distributed mainly in the magnocellular PVN. Double in situ hybridization histochemistry revealed that some of the iNOS- (48.4%) or nNOS-positive cells (34. 3%) in the parvocellular PVN expressed CRF mRNA. The results demonstrate that LPS-induced sepsis causes significant increases in nNOS and iNOS gene expression with different time-courses and distributions, and that iNOS mRNA was more frequently co-localized with CRF-producing parvocellular neurons in the PVN. Thus, NO produced by iNOS and nNOS may play an important role in the neuroendocrine response to an immune challenge. Distinct differences in the distribution and time-course changes of iNOS and nNOS suggest different roles for the hypothalamic-pituitary-adrenal axis and/or neurohypophyseal system.
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PMID:Distinct distribution and time-course changes in neuronal nitric oxide synthase and inducible NOS in the paraventricular nucleus following lipopolysaccharide injection. 1006 18

We investigated the enzymes involved in the NADPH-diaphorase (d) reaction in the rat and pig bladder urothelium. The urothelial cell layer displayed intense and uniform NADPH-d activity. Preincubation with the flavoprotein inhibitor diphenyleneiodionium chloride (DPI) and the alkaline phosphatase inhibitor levamisole concentration-dependently decreased the urothelial NADPH-d activity. Immunoreactivities to neuronal (n), endothelial (e), or inducible (i) nitric oxide synthase (NOS) were not detected in rat or pig urothelial cells. In rats, the urothelium was uniformly immunoreactive for NADPH cytochrome P450 reductase, whereas the pig urothelium displayed inconsistent labeling. In lipopolysaccharide (LPS)-treated rats, the bladder urothelium showed positive iNOS immunoreactivity. The iNOS labeling was found predominantly in cells located in the basal layer of the urothelium. In the pig bladder mucosa, a Ca2+-dependent NOS activity was evident in cytosolic and particulate fractions that was quantitatively comparable to the NOS activity found in the smooth muscle. In ultrastructural studies of urothelial cells, NADPH-d reaction products were found predominantly on membranes of the nuclear envelope, endoplasmatic reticulum and mitochondria. In conclusion, NADPH-d staining of the urothelium cannot be taken as an indicator for the presence of constitutively expressed NOS. Activity of alkaline phosphatase and cytochrome P450 reductase may account for part of the NADPH-d reaction in urothelial cells. However, LPS treatment of rats caused expression of iNOS in urothelial cells.
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PMID:Morphological and biochemical investigation of nitric oxide synthase and related enzymes in the rat and pig urothelium. 1033 Apr 50

In contrast to the vertebrate immune system, nearly nothing is known about the immunological role of nitric oxide (NO) in invertebrates. This study provides evidence of the presence of a NO synthase (NOS) activity in an immune-competent, macrophage-like insect hemocyte line, previously established from larvae of the lepidopteran insect Estigmene acraea. As proven by photometric determination of nitroblue tetrazolium reduction after cell fixation, the E. acraea cells possess NADPH diaphorase (NADPHd) activity. This NADPH diaphorase activity was NADPH dependent, not inhibitable by superoxide dismutase, influenced by extracellular addition of L-arginine, and inhibited in a dose-dependent manner by the specific NOS inhibitor Nomega-monomethyl-L-arginine. Furthermore, the NADPH diaphorase activity was stimulated within 30 min by the addition of insect pathogenic bacteria (Bacillus thuringiensis var. kurstaki, Photorhabdus luminescens), bacterial lipopolysaccharide, and silica beads. In activated E. acraea cell suspensions strongly increased amounts of L-citrulline and enhanced levels of total nitrite/nitrate (as NO derivates) can be determined. This is the first report on stimulable NOS activity in insect hemocytes.
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PMID:Stimulation of NO synthase activity in the immune-competent lepidopteran Estigmene acraea hemocyte line. 1036 82

We previously described a long-lasting overproduction of nitric oxide (NO) in cirrhotic patients with spontaneous bacterial peritonitis. The aim of the present study was to investigate the presence of the inducible NO pathway in peritoneal macrophages. Ascitic fluids were collected from 29 patients with cirrhosis, aged between 35 and 82 years. Peritoneal macrophages were isolated and cultured in the presence or absence of 1 microg/ml lipopolysaccharide and/or 500 units/ml interferon-gamma (IFN-gamma) for 6 days. NO production was measured as nitrate+nitrite (NO(x)), inducible NO synthase (iNOS) protein expression was analysed by immunocytochemistry and Western blot analysis using a specific anti-(human iNOS) antibody, and the catalytic activity of NOS was revealed by cytochemical staining for NADPH-dependent diaphorase. Cultured macrophages spontaneously released small amounts of NO(x) [median (10-90th percentile) of 18 separate experiments: 3.3 (0-8) micromol/l]. Addition of lipopolysaccharide alone or in combination with IFN-gamma to the culture medium did not change the levels of NO(x), while IFN-gamma alone dramatically increased NO production [13.4 (3.5-28.3) micromol/l; P<0.001]. Macrophages were stimulated by IFN-gamma to a greater extent in patients with recent spontaneous bacterial peritonitis (n=13) than in those in a stable clinical condition (n=18) [19.8 (10.5-30.1) and 10.0 (3.2-14.5) micromol/l respectively; P<0.001]. Macrophages freshly isolated or stimulated with IFN-gamma expressed iNOS protein, as shown by Western blot and immunocytochemical analysis, and stained for NADPH diaphorase. Our findings demonstrate the presence of iNOS protein in peritoneal macrophages from cirrhotic patients. The role of IFN-gamma appears to be a determinant for the up-regulation of NO production, particularly under conditions of infection. Therefore peritoneal macrophages producing large amounts of NO at the site of infection may contribute to maintaining splanchnic vasodilation in these patients.
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PMID:Up-regulation of nitric oxide production by interferon-gamma in cultured peritoneal macrophages from patients with cirrhosis. 1049 39

We have obtained evidence that co-incubation of thioglycollate-elicited peritoneal macrophages with an aqueous extract of Tinospora tuberculata inhibits lipopolysaccharide-stimulated excessive production of nitric oxide (NO) in vitro. This effect is concentration-dependent and appears to involve suppression of both inducible nitric oxide synthase (iNOS) activity and NADPH-diaphorase activity, thus altering NO production. As NO is one of the critical mediators in various disorders and iNOS inhibitors may have therapeutic potential, these results may explain some aspects of the multifunctional properties of Tinospora tuberculata, which has been used in various folk remedies in southeast Asia and China.
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PMID:Tinospora tuberculata suppresses nitric oxide synthesis in mouse macrophages. 1074 61


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