UNIPROT:P43026 - FACTA Search

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Query: UNIPROT:P43026 (lipopolysaccharide)
62,215 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. The induction of cyclo-oxygenase-2 (COX-2) afforded by bacterial lipopolysaccharide (LPS, endotoxin) in bovine aortic endothelial cells (BAEC) is mediated by tyrosine kinase. LPS also causes the generation of several cytokines including interleukin-1 beta (IL-1 beta), tumour necrosis factor-alpha (TNF-alpha), epidermal growth factor (EGF) and platelet-derived growth factor (PDGF). This study investigates whether endogenous IL-1 beta, TNF-alpha, EGF or PDGF contribute to the induction of COX-2 elicited by LPS in BAEC and if their action is due to activation of tyrosine kinase. Furthermore, we have studied the induction of COX-2 by exogenous cytokines. 2. Accumulation of 6-oxo-prostaglandin (PG) F1 alpha in cultures of BAEC was measured by radioimmunoassay at 24 h after addition of either LPS (1 microgram ml-1) alone or LPS together with a polyclonal antibody to one of the various cytokines. In experiments designed to measure 'COX activity', 6-oxo-PGF1 alpha generated by BAEC activated with recombinant human IL-1 beta, TNF-alpha, EGF or PDGF for 12 h was measured after incubation of washed cells with exogenous arachidonic acid (30 microM for 15 min). Western blot analysis determined the expression of COX-2 protein in BAEC. 3. The accumulation of 6-oxo-PGF1 alpha caused by LPS in BAEC was attenuated by co-incubation with one of the polyclonal antibodies, anti-IL-1 beta, anti-TNF-alpha, anti-EGF, anti-PDGF or with the IL-1 receptor antagonist, in a dose-dependent manner. Exogenous IL-1 beta, TNF-alpha or EGF also caused an increase in COX activity, while PDGF was ineffective. The increase in COX activity elicited by IL-1,beta(10 ng ml-1), TNF-alpha (100 ng ml-1) or EGF (1000 ng ml-1) in BAEC was attenuated by erbstatin (0.005 to 5 microg ml-1), as was the expression of COX-2 protein measured by Western blot analysis.4. PDGF (10 ng ml-1) significantly augmented the rise in COX activity and COX-2 protein caused by shorter incubation of BAEC with LPS (1 microg ml-1 for 3 h). Combination of PDGF (10 ng ml-1) with a low concentration of IL-l beta (1 ng ml-1) for 12 h, also increased 'COX activity', but combination of PDGF and TNF-alpha (10 ng ml-1) did not show any increased activity.5. These results suggest that (i) the induction of COX activity and COX-2 protein elicited by LPS in BAEC is mediated by TNF-alpha with lesser contributions from PDGF, EGF or IL-1 beta; (ii) exogenous IL-1 beta,TNF-alpha or EGF alone induce COX-2 activity and protein in BAEC; (iii) PDGF synergizes with IL-1 beta,but not TNF-alpha, to cause expression of COX-2; and (iv) the induction of COX-2 protein and activity caused by these cytokines involves the activation of tyrosine kinase.
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PMID:Cytokine-mediated induction of cyclo-oxygenase-2 by activation of tyrosine kinase in bovine endothelial cells stimulated by bacterial lipopolysaccharide. 758 49

Insect hemocytes (blood cells) synthesize the major nonself recognition protein (47 kDa) during 3rd instar larvae (V.J. Marmaras, S. Tsakas, Dev. Biol. 129, 294-303 (1988)). In this study we show the presence of the 47 kDa protein in plasmatocytes (main hemocyte type) and prohemocytes. In plasmatocytes this protein appears to be localized both in vesicles and in the cell surface. The cell surface-associated 47 kDa protein was released from membrane fraction by 1 M NaCl, indicating that it is not tightly bound. Bacterial lipopolysaccharide (LPS) can function on isolated hemocytes from Ceratitis capitata larvae, inducing their spreading and degranulation. During degranulation (exocytosis) the plasmatocytes release the 47 kDa protein, among others. This protein could not be normally traced in serum, nor is it released by basal secretion. The secretion of the 47 kDa protein was found to be LPS-dependent, whereas its presence on plasmatocyte surface is LPS independent. LPS-stimulated exocytosis of the 47 kDa protein appears to be dependent on protein tyrosine phosphorylation. We have now demonstrated that LPS increases tyrosine phosphorylation of 19 and 22 kDa polypeptides in C. capitata hemocytes. Inhibition of the LPS-induced tyrosine phosphorylation mediated by tyrosine kinase inhibitor, genistein, was accompanied by the inhibition of the secretion of the 47 kDa protein. These results support the hypothesis that tyrosine protein phosphorylation is a signal reaction in hemocytes after LPS exposure. These LPS responses of insect plasmatocytes show strong similarities to mammalian macrophages (S. Weinstein et al., J. Immunol. 151, 3829-3838 (1993)). In a model we propose that the LPS-independent cell surface-associated 47 kDa protein is responsible for the phagocytosis and for the formation of nodules and capsules, whereas the LPS-dependent secreting counterpart is responsible for the extracellular killing of bacteria.
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PMID:Lipopolysaccharide-stimulated exocytosis of nonself recognition protein from insect hemocytes depend on protein tyrosine phosphorylation. 764 28

The addition of fMet-Leu-Phe or phorbol 12-myristate 13-acetate to human neutrophils stimulates phospholipase D activity as evidenced by the release of phosphatidic acid and the generation of diacylglycerol, and in the presence of ethanol the formation of phosphatidyl ethanol. The activation of phospholipase D by either the chemotactic factor or active phorbol ester is inhibited by the tyrosine kinase inhibitor erbstatin. The fMet-Leu-Phe-induced stimulation of this enzyme is greatly potentiated in cells which have been preincubated with low concentrations of lipopolysaccharide and serum. The presence of serum is essential for the potentiation by low concentrations of lipopolysaccharide. Moreover, the monoclonal antibody MY4(IgG2b) against CD14 inhibits the potentiation by the low concentration of lipopolysaccharide. These data suggest three important points. First, a tyrosine kinase step is necessary for the activation of phospholipase D. This suggests that the phospholipase D enzyme needs to be phosphorylated on tyrosine residues to be activated. Second, low concentrations of lipopolysaccharide, in the presence of serum, can potentiate the stimulated activity of this enzyme. Third, the priming action of the lipopolysaccharide-serum complex is mediated by CD14.
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PMID:Lipopolysaccharide in combination with serum potentiates the stimulated activity of phospholipase D in human neutrophils via CD14. 768 89

CD14, a glycosylphosphatidylinositol-anchored glycoprotein of leukocytes, binds endotoxin (lipopolysaccharide (LPS)) with high affinity. After the murine pre-B cell line 70Z/3 is transfected with DNA encoding human CD14 (hCD14), the resultant stably transfected cell line, 70Z/3-hCD14, responds to 1000-fold lower LPS concentrations than the parental CD14-negative line. We have used 70Z/3-hCD14 cells, RAW264.7 cells, and elicited murine peritoneal exudate macrophages (PEM) to study LPS-induced protein tyrosine phosphorylation. LPS induces the rapid tyrosine phosphorylation of a 38-kDa protein (p38) in 70Z/3-hCD14 cells, PEM, and RAW264.7 cells and of two isoforms of mitogen-activated protein kinases (MAPK) in only RAW264.7 cells and PEM. p38 can be distinguished from the MAPK isoforms based on differences in mobilities on SDS-polyacrylamide gel electrophoresis and the lack of reactivity of p38 with anti-MAPK antibody even after dephosphorylation with potato acid phosphatase. Synthetic lipid A induces p38 phosphorylation in 70Z/3-hCD14 cells, whereas phorbol 12-myristate 13-acetate and interferon-gamma fail to induce tyrosine phosphorylation of p38. Pretreatment of 70Z/3-hCD14 cells with anti-hCD14 monoclonal antibody or the tyrosine kinase inhibitor herbimycin A inhibits LPS-induced tyrosine phosphorylation of p38. These results suggest that increased protein tyrosine phosphorylation occurs rapidly after LPS binds to CD14 and is likely to be an important event in mediating LPS-induced cell activation.
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PMID:Endotoxin induces rapid protein tyrosine phosphorylation in 70Z/3 cells expressing CD14. 769 11

The expression of many genes is altered upon the activation of macrophages by bacterial LPS. These genes play a crucial role in the orchestration of various responses to protect the host against infection. A novel 2.3 kilobase (kb) cDNA, designated IRG1, was obtained from a cDNA library prepared with RNA isolated from RAW 264.7 following lipopolysaccharide stimulation. Sequence analysis of the clone revealed no identity to any known genes but showed the presence of many potential phosphorylation sites suggesting that IRG1 protein product may be regulated at this level. Furthermore, IRG1 contains the motif for glycosaminoglycan attachment site, implying that IRG1 may be a proteoglycan. By interspecific back-cross analysis, Irg1 was mapped to mouse chromosome 14 linked to Tyrp2 and Rap2a. The IRG1 message appears 1.5 h following LPS exposure and its induction was not dependent on new protein synthesis. In fact, cycloheximide induced the expression of IRG1, suggesting that a protein repressor prevents the expression of IRG1 when uninduced. The role of the protein kinase A pathway in regulating the induction of IRG1 by LPS is questionable, because although forskolin inhibited its induction, neither dibutyrl-cAMP nor 8-(4-chlorophenylthio)-cAMP had much effect on its expression. In contrast, activation of protein kinase C potentiated the LPS response. Chelation of extracellular calcium inhibited IRG1 4 h after LPS induction, while increasing intracellular calcium had little effect on the levels of the IRG1 transcript. Inhibiting tyrosine phosphorylation abrogated the induction of IRG1 by LPS. Hence, the induction of IRG1 by LPS is mediated by tyrosine kinase and protein kinase C pathway.
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PMID:Cloning and analysis of gene regulation of a novel LPS-inducible cDNA. 772 48

The production and the effects of tumor necrosis factor alpha (TNF alpha) have been studied in isolated glomeruli and cultured glomerular cells from animal or human origin. Glomeruli from rats injected with E. coli lipopolysaccharide (LPS) and glomeruli exposed to LPS in vitro release TNF alpha into the medium. The glomerular cells responsible for TNF alpha synthesis are mesangial cells. Production of TNF alpha is controlled by other locally produced mediators. Prostaglandin E2 and interleukin-10 are inhibitory. Hydroxyl radicals stimulate the transformation of the transmembrane form of TNF alpha into its soluble form which is secreted into the medium. TNF alpha acts on its producing cells and on the neighbouring cells. It induces contraction of mesangial cells, increases the synthesis of a variety of local mediators including prostaglandins, platelet-activating factor and chemotactic agents, particularly interleukin-8. Synthesis of this cytokine implies activation of tyrosine kinase and of transcription factors, essentially NFkB. Taken together, these results suggest that TNF alpha plays a marked role in the development of glomerular injury and incite us to search for treatments inhibiting its synthesis and its effects.
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PMID:[Production and proinflammatory activity of tumor necrosis factor alpha in the glomerulus]. 778 39

Nuclear factor kappa B (NF-kappa B), consisting of p50 and p65, is bound to a cytoplasmic retention protein, I kappa B, in a resting state, and the stimulation of cells with a variety of inflammatory stimuli induces the dissociation of NF-kappa B from I kappa B and the nuclear translocation of NF-kappa B, thereby activating several genes involved in inflammatory responses, such as interleukin (IL)-6, IL-8, and tumor necrosis factor alpha. In order to elucidate the precise mechanism of NF-kappa B activation, we have established lipopolysaccharide (LPS)-dependent NF-kappa B activation in a cell-free system using plasma membrane-enriched, cytosol, and nuclear fractions extracted from a human monocytic cell line, THP-1, by disruption with sonication followed by a differential centrifugation. The combination of plasma membrane-enriched fraction and cytosol was sufficient to activate NF-kappa B in a LPS/CD14-dependent manner only in the presence of ATP as judged by the binding of NF-kappa B to the IL-8 gene kappa B site on an electrophoretic mobility shift assay. LPS-dependent NF-kappa B activation was inhibited by protein kinase inhibitors, such as staurosporine, herbimycin A, tyrphostin, and genistein, but not mitogen-activated protein kinase substrate, cGMP-dependent protein kinase, cAMP-dependent protein kinase, protein kinase C, and calmodulin-dependent protein kinase II inhibitory peptides, suggesting that staurosporine-sensitive kinase(s) as well as tyrosine kinase(s) are involved in LPS-mediated NF-kappa B activation. In addition, LPS induced the phosphorylation of I kappa B-alpha, starting at 5 min after the stimulation in a cell-free system. Moreover, the phosphorylation was inhibited by herbimycin A and tyrphostin, but not staurosporine, suggesting that these protein kinase inhibitors act at distinct steps of signal transmission. Establishment of ligand-dependent activation of NF-kappa B in a cell-free system will facilitate identification of protein kinase(s) and its substrate(s) involved in LPS-mediated NF-kappa B activation.
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PMID:Establishment of lipopolysaccharide-dependent nuclear factor kappa B activation in a cell-free system. 787 68

Two isoforms of cyclooxygenase (COX) have been identified in eukaryotic cells: a constitutively expressed COX-1 and mitogen-inducible COX-2, which is selectively expressed in response to various inflammatory stimuli. Thus, COX-2 instead of COX-1 is implicated to produce prostanoids mediating inflammatory responses. Major efforts have been focused on identifying nonsteroidal anti-inflammatory drugs (NSAIDS) which can selectively inhibit the enzyme activity of COX-2. Such NSAIDS would be more desirable anti-inflammatory agents in comparison to NSAIDS which inhibit both COX-1 and COX-2. Other than glucocorticoids, pharmacological agents which can selectively suppress the expression of COX-2 without affecting that of COX-1 have not been identified. We report here that radicicol, a fungal antibiotic, is a potent protein tyrosine kinase inhibitor, and that it inhibits the expression of COX-2 without affecting COX-1 expression in lipopolysaccharide (LPS)-stimulated macrophages with the IC50 value of 27 nM. Radicicol inhibited tyrosine phosphorylation of p53/56lyn, a Src family tyrosine kinase and one of the major tyrosine-phosphorylated proteins in LPS-stimulated macrophages. Radicicol also inhibited COX-2 expression in vivo in glomeruli of rats with experimental glomerulonephritis induced by the anti-glomerular basement membrane antibodies, in which COX-2 expression is known to be enhanced. The enzyme activity of COX-1 or COX-2 was not affected by radicicol in macrophages. Radiciciol also suppressed the COX-2 expression induced by IL-1 beta in rat smooth muscle cells. Other protein tyrosine kinase inhibitors suppressed the LPS-induced COX-2 expression in macrophages but at much higher concentrations than needed for radicicol. Radicicol did not inhibit the COX-2 expression induced by phorbol 12-myristate 13-acetate in macrophages. These results suggest that the activation of tyrosine-specific protein kinases is the proximal obligatory step in the LPS-induced signal transduction pathway leading to the induction of COX-2 expression in macrophages. The magnitude of the inhibition of COX-2 protein synthesis by radicicol was much greater than that of the steady state levels of COX-2 mRNA. These results suggest that radicicol inhibits COX-2 expression mainly at post-transcriptional steps.
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PMID:Radicicol, a protein tyrosine kinase inhibitor, suppresses the expression of mitogen-inducible cyclooxygenase in macrophages stimulated with lipopolysaccharide and in experimental glomerulonephritis. 789 Jun 56

Activation of monocytes by bacterial lipopolysaccharides (LPSs) is a central component in the pathogenesis of septic shock syndrome. Interleukin 10 (IL-10) is a potent monocyte-deactivating factor and transcriptionally inhibits LPS-induced expression of proinflammatory mediators. The intracellular signaling pathways of LPS have been only partially characterized and mechanisms of IL-10 signaling remain unknown. We show that LPS activates the protein tyrosine kinase (PTK) p56lyn and that this is associated with tyrosine phosphorylation of the protooncogene product Vav. These events are completely blocked by the tyrosine kinase inhibitor herbimycin A. LPS also increases Ras activation in monocytes. LPS-triggered phosphorylation of mitogen-activated protein kinase is a downstream activation event that is also reduced by herbimycin A. Analysis of the IL-10 effects shows that it completely inhibits the p56lyn tyrosine kinase activation and all other subsequent events in this pathway including Ras activation. The IL-10 effects are selective since it reduced PTK-dependent cytokine mRNA expression but not the PTK independent induction of c-jun and c-fos mRNA in LPS-activated monocytes. These results identify the Ras signaling pathway as a component of intracellular signaling in LPS-stimulated monocytes and define early events in this response as targets of monocyte deactivation by IL-10.
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PMID:Monocyte deactivation by interleukin 10 via inhibition of tyrosine kinase activity and the Ras signaling pathway. 807 29

Nitric oxide (NO) formation via the expression of an endotoxin- and cytokine-inducible NO synthase (iNOS) within the vascular smooth muscle is thought to be responsible for the cardiovascular collapse that occurs during septic shock and antitumor therapy with cytokines. Because the molecular mechanisms that underlie induction of iNOS are still unclear and because tyrosine kinases are implicated in interleukin-1 beta (IL-1 beta)-induced prostaglandin synthesis in mesangial cells and in NO generation by an insulinoma cell line, we investigated the influence of tyrosine kinase inhibitors on iNOS induction in cultured rat aortic smooth muscle cells (RASMC). The production of biologically active NO was demonstrated by L-arginine-dependent guanosine 3',5'-cyclic monophosphate (cGMP) accumulation after a 3-h exposure to either IL-1 beta or lipopolysaccharide (LPS). Pretreatment of RASMC for 30 min with the tyrosine kinase inhibitor genistein prevented both IL-1 beta- and LPS-elicited cGMP accumulation in a concentration-dependent manner. Geldanamycin, a chemically different tyrosine kinase inhibitor, also blocked cGMP formation in response to both LPS and IL-1 beta at nanomolar concentrations. Genistein and geldanamycin inhibited cGMP accumulation even when added 90 min after LPS exposure, but no inhibition was observed when they were included at later time points (120-180 min), suggesting that the inhibitors had no direct effect on iNOS activity after its induction. Formation of cGMP in response to sodium nitroprusside and to NO released from bovine aortic endothelial cells remained virtually unaffected by genistein and geldanamycin.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Tyrosine kinase inhibitors suppress endotoxin- and IL-1 beta-induced NO synthesis in aortic smooth muscle cells. 821 7

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