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)

Maternal infection is a cause of spontaneous abortion and preterm labor in humans, but the pathophysiology is unclear. We hypothesized that eicosanoids play an important role in infection-driven pregnancy loss. To investigate this hypothesis, we administered lipopolysaccharide (LPS) to pregnant C3H/HeN mice and found that LPS administration caused fetal death in a dose-dependent fashion. Pretreatment with indomethacin significantly decreased the proportion of fetal death from 83% to < 25% in mice injected with 10 micrograms of LPS. Also, decidual explants from LPS-treated mice produced significantly more inflammatory eicosanoids, including prostaglandins E2 and F2 alpha and thromboxane B2, than controls. We investigated the regulatory mechanisms responsible for increased decidual prostanoid production in response to LPS. Western and Northern blots demonstrated that decidual protein and mRNA levels of a recently recognized highly inducible form of cyclooxygenase, COX-2, were substantially increased in mice treated with LPS. Induction of COX-2 was rapid: mRNA was detected 30 min after LPS injection. In contrast, another form of cyclooxygenase, COX-1, was only minimally induced in response to LPS. Our data indicate that LPS induces decidual prostanoid production via increased COX-2 expression. Since LPS-mediated fetal death is markedly diminished by pretreatment with indomethacin, COX-2-mediated eicosanoid production is likely a key pathophysiologic event in LPS-mediated fetal death.
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PMID:Bacterial lipopolysaccharide-mediated fetal death. Production of a newly recognized form of inducible cyclooxygenase (COX-2) in murine decidua in response to lipopolysaccharide. 786 Jul 53

Prostaglandin synthesis represents one means by which macrophages modulate inflammation. The initial enzyme in the metabolism of arachidonic acid to prostaglandins is cyclooxygenase (COX). Both constitutive (COX-1) and inducible (COX-2) isoforms are recognized. We previously showed that COX activity of rat peritoneal macrophages (PM) exceeds that of alveolar macrophages (AM). In this study, we correlated the steady-state levels of COX-1 and COX-2 proteins with COX activity in resident AM and PM. Freshly obtained AM contained lower levels of COX-1 than did fresh PM. Neither contained substantial amounts of COX-2 in the basal state, but both cell types demonstrated induction when cultured with lipopolysaccharide; once again, COX-2 levels in PM exceeded those in AM. Despite COX-2 induction under these circumstances, its contribution to prostaglandin production appeared to be modest. We conclude that, although both isoforms of COX are expressed in rat AM and PM, COX-1 is responsible for the majority of enzyme activity in both the basal and stimulated states. The lesser prostaglandin synthetic capacity of AM than of PM appears to be the consequence of lower steady-state levels of both COX proteins.
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PMID:Expression and role of cyclooxygenase isoforms in alveolar and peritoneal macrophages. 786 49

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

Cyclo-oxygenase, a key enzyme in prostaglandin production, is mainly in the constitutive form (COX-1) in gastric mucosa, whereas leucocytes have an inducible enzyme (COX-2). We report that indomethacin and its glycolic acid ester acemetacin have quantitatively different effects on prostanoid synthesis in these tissues. Human gastric mucosa (fresh operation specimens, cut finely and washed), and 1-1.5 x 10(6) human blood leucocytes stimulated with lipopolysaccharide (5 micrograms/ml), were incubated with acemetacin or indomethacin (0.1, 1, 10, 100 micrograms/ml). Eicosanoids in the medium were measured by radioimmunoassay (RIA). In leucocytes, acemetacin and indomethacin were more potent cyclo-oxygenase inhibitors than in the gastric mucosa, and both reduced the PGE levels to similar extents (70-98% and 72-100% respectively, n = 5). LTB4 was reduced only at 100 micrograms/ml of either drug. In gastric mucosal incubates, acemetacin was less potent than indomethacin in causing a concentration-related inhibition of PGE accumulation (19-74% vs 34-84%; n = 6, p < 0.05). Acemetacin was also less potent than indomethacin in reducing gastric 6-keto-PGF1 alpha and TXB2 (by 11-79% vs 45-72%, and 0-80% vs 29-80% respectively, p < 0.05). The finding that acemetacin is equipotent to indomethacin on leucocyte cyclo-oxygenase (inducible enzyme, COX-2) but less active on the gastric mucosa (COX-1) is consistent with an effective analgesic and anti-inflammatory activity of acemetacin coupled with better gastric tolerance than that to indomethacin.
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PMID:Acemetacin and indomethacin: differential inhibition of constitutive and inducible cyclo-oxygenases in human gastric mucosa and leucocytes. 814 13

We investigated regulation of macrophage prostaglandin production during activation by interferon gamma (IFN-gamma) and lipopolysaccharide (LPS). An in vitro model was established using the mouse macrophage-like cell line RAW 264.7. Cells were cultivated in the presence of IFN-gamma and LPS for up to 48 h and changes in the secretion of nitric oxide (NO.) and tumor necrosis factor alpha (TNF-alpha) were observed as activation markers. Under these conditions a prompt and strong increase in PGE2 production was found in the first 8 h followed by nearly constant generation of PGE2 during the next 40 h. In contrast, the activity of prostaglandin endoperoxide synthase (PGHS), measured as PGE2 production of microsomal protein fractions, was also increased, but reached a clear maximum at 24 h. Recently a second form of PGHS was cloned (PGHS-2) and specific antibodies and mRNA probes for both isoforms are available. PGHS-2 enzyme was expressed maximally after 24 h of activation whereas PGHS-1 was not influenced. In the presence of IFN-gamma and LPS, PGHS-2 mRNA expression reached a maximum at 8 h but PGHS-1 mRNA was not induced during the whole time period. These data indicate that changes in PG synthesis following macrophage activation are due to regulation of PGHS-2 expression.
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PMID:Transient expression of prostaglandin endoperoxide synthase-2 during mouse macrophage activation. 814 18

Two isoforms of cyclooxygenase (COX) have been identified in eukaryotic cells: COX-1 encoded by a 2.8-kb mRNA, and a mitogen-inducible COX-2 encoded by a 4-kb mRNA. We have cloned the COX-1 and COX-2 cDNAs from the cDNA library constructed from lipopolysaccharide (LPS)-stimulated rat peritoneal macrophages. The deduced amino acid sequence showed that COX-1 contained 602 amino acids, whereas COX-2 contained 604 amino acids. There is 95% conservation of the nucleotide sequence in the open reading frame of COX-1 between the rat and the mouse, while the homology of the 3' untranslated region is 68% except for a 150 bp segment adjacent to the stop codon which is nonhomologous with the mouse. Transfection of both COX cDNAs into Cos-7 cells resulted in increased COX activity. In rat vascular smooth muscle cells, interleukin-1 beta selectively increased the expression of COX-2, but not that of COX-1, as assessed by enzyme activity, immunoprecipitation of COX proteins, and mRNA analysis. Only the brain among tissues tested exhibits basal expression of COX-2 as the major form of the enzyme. However, COX-2 mRNA was expressed in vivo in the lung and kidney, but not in the heart, after systemic administration of LPS, suggesting that COX-2 but not COX-1 plays a major role in producing COX-derived products of arachidonic acid during endotoxic shock. Thus, the two COX isoforms were differentially expressed, and COX-2 was selectively induced in response to inflammatory stimuli in rats.
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PMID:Cloning two isoforms of rat cyclooxygenase: differential regulation of their expression. 827 23

Previous studies from our laboratory suggested that phorbol 12-myristate 13-acetate (TPA) stimulates prostaglandin E2 (PGE2) production by inducing de novo synthesis of prostaglandin H synthase (PHS) in a rat tracheal cell line. We report here an extension of this work to further elucidate the mechanisms by which TPA (and epidermal growth factor) stimulates PGE2 production. We used the rat tracheal cell line EGV6, which has a lower basal level of PGE2 production and responds to TPA and EGF stimulation with a much greater increase in PGE2 synthesis than the previously used cell line, Incubation of EGV6 cultures with TPA or EGF resulted in a time- and dose-dependent increase in PGE2 synthesis up to 40-fold and 6-fold, respectively. Serum also stimulated PGE2 synthesis, while bombesin, retinoic acid, and bacterial lipopolysaccharide did not. PHS protein levels in microsomal preparations from the cells were estimated by Western analysis. Antibodies raised against murine PHS-2 cross reacted with the EGV-6 PHS while several antibody preparations that react with PHS-1 from ram or mouse reacted poorly with the cellular preparation. TPA treatment increased the de novo synthesis of PHS-2 while dexamethasone treatment reduced the response to TPA. Northern blot analysis of mRNA from EGV6 cultures using a ram PHS cDNA revealed a 2.8- and a 4.5- to 4.9-kb (designated 4.9 kb) transcript. Treatment with TPA or EGF increased the expression of both transcripts and this effect was further enhanced by cyclohexamide. To further define the PHS mRNA species of EGV6 cells, two well-characterized murine PHS cDNA probes were used. The constitutive murine PHS cDNA probe hybridized only with the 2.8-kb transcript, and the inducible murine PHS cDNA hybridized only with the 4.9-kb transcript. The rates of induction as well as degradation of the 4.9-kb PHS mRNA were much more rapid than those of the 2.8-kb mRNA species. Dexamethasone partially inhibited the induction of both PHS transcripts by TPA. Southern analysis of genomic EGV6 DNA indicated the presence of two distinct PHS genes in these cells. Taken together these findings indicate that two PHS genes are expressed in rat tracheal epithelial cells. In contrast to the PHS genes expressed in murine (and chicken) fibroblasts in which only the gene coding for the larger mRNA species is transcriptionally regulated, in the rat tracheal cells both genes are positively regulated by TPA and EGF and downregulated by glucocorticoids.
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PMID:Phorbol ester and epidermal growth factor enhance the expression of two inducible prostaglandin H synthase genes in rat tracheal epithelial cells. 832 87

Prostaglandin endoperoxide synthase (PHS) catalyzes the committed step in the biosynthesis of prostaglandins and thromboxane. We recently observed dissociation of PHS activity and enzyme mass measured in an immunoassay of endothelial cells exposed to tumor necrosis factor. These data and observations by others suggested that endothelial cells express an alternate PHS. We now report the molecular cloning of human PHS type II from an endothelial cell cDNA library. The protein encoded by this cDNA shares 61% identity with the human PHS I. Southern analysis demonstrated a single copy of PHS II and we found a polymorphism in approximately 5% of the population. PHS II mapped to chromosome 1, in contrast to PHS I, which is on chromosome 9. The PHS II cDNA hybridized strongly to a 4.3-kilobase (kb) message from endothelial cells. Stimulation of the cells with tumor necrosis factor, phorbol 12-myristate 13-acetate, lipopolysaccharide, or interleukin-1 increased mRNA levels for PHS II, and this change correlated well with increased prostacyclin biosynthesis. Cycloheximide induced PHS II mRNA without a corresponding activity increase demonstrating that translation of the 4.3-kb message is required for increased prostacyclin biosynthesis. We conclude that expression of PHS II may have important pathophysiological effects in the vasculature.
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PMID:Molecular cloning of human prostaglandin endoperoxide synthase type II and demonstration of expression in response to cytokines. 847 46

Meloxicam is a new nonsteroidal anti-inflammatory drug (NSAID) derived from enolic acid. Meloxicam has shown potent anti-inflammatory activity in animal models together with low gastrointestinal and renal toxicity. Studies were undertaken to compare meloxicam to other NSAIDS in their ability to inhibit either constitutive cyclooxygenase (COX-1) or inducible cyclooxygenase (COX-2). COX-1 was isolated as a cell-free enzyme from bovine seminal vesicles or bovine brain or was present in nonstimulated macrophages derived from the guinea-pig peritoneum. COX-2 was induced in peritoneal macrophages stimulated by lipopolysaccharide (LPS) or isolated as a cell-free enzyme from sheep placenta. Of all NSAIDs tested, meloxicam was the most selective inhibitor of COX-2 in intact cells. In cell-free enzyme preparations, however, meloxicam showed the same activity against COX-1 and COX-2. All other NSAIDs tested were more potent inhibitors of COX-1 than of COX-2. The inducible cyclooxygenase COX-2 has been implicated in the mediation of the inflammatory reaction, whereas the products of the constitutive cyclooxygenase COX-1 have cytoprotective effects in the gastric mucosa, support microcirculation in the kidney, and are antithrombogenic. Therefore, differential inhibitory effects of NSAIDs on COX-1 and COX-2 may have a bearing on the risk-benefit profile displayed in clinical practice. Meloxicam shows a preferential inhibitory effect on COX-2 over COX-1, which may be directly related to the favorable tolerability profile with potent anti-inflammatory effects observed in animal studies.
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PMID:Meloxicam: influence on arachidonic acid metabolism. Part 1. In vitro findings. 853 64

Prostaglandins are pro-inflammatory but are gastroprotective. The gastric mucosa synthesizes prostaglandins mainly via constitutive cyclooxygenase (COX-1), whereas leucocytes have inducible enzyme (COX-2). Nimesulide (CAS 51803-78-2) differentially inhibited prostanoid synthesis in these human tissues as well as with in vitro enzyme assays, and was less potent than indometacin (CAS 53-86-1) on COX-1. Fresh human gastric mucosa was cut finely, washed and pre-incubated (100 mg in 1 ml phosphate buffered saline pH 7.4) with or without nimesulide or indometacin (0.1-100 micrograms/ml; 0 degree C; 30 min). The fluid was replaced with fresh identical solution, incubated (37 degrees C; 30 min) and the solution assayed. Isolated leucocytes from human peripheral blood were incubated (1-1.5 x 10(6), 2 ml Krebs' solution) with or without nimesulide or indometacin (0.1-100 micrograms/ml; 37 degrees C; 1 h), stimulated with lipopolysaccharide (5 micrograms/ml), further incubated for 24 h at 37 degrees C and the medium assayed for the prostanoids PGE, TXB2, 6-keto-PGF1 alpha and the leukotriene LTB4 by radioimmunoassay (RIA). In vitro assays with COX-1 from ram seminal vesicles, or COX-2 from sheep placenta, were performed by pre-incubating the enzymes with vehicle alone (controls) or with drug for 5 min at 37 degrees C. Arachidonate (10 mumol/l) was added and further incubated for 2 min at 37 degrees C. Reactions were terminated and PGE determined by RIA. Both drugs caused concentration-related inhibitions of prostanoid accumulation in incubates of both tissues. Nimesulide reduced PGE accumulation more potently in incubates of stimulated leucocytes than of gastric mucosa. With gastric tissue, nimesulide was less potent than indometacin by approximately 6-22 fold (IC50 for PGE, TXB2, 6-keto-PGF1 alpha, respectively; 14.8 vs 2.5; 12.8 vs 1.0; 31.1 vs 1.4 mumol/l; p < 0.05 to 0.02). With the leucocytes, the concentrations of both drugs, particularly indometacin were not low enough to calculate the IC50. With the in vitro assay, nimesulide (0.01 to 100 mumol/l) did not inhibit PGE formation by COX-1 but caused a concentration-related inhibition of PGE formation by COX-2 (4-60%). These results are consistent with the effective analgesic/anti-inflammatory activity of nimesulide coupled with better gastric tolerance compared to indometacin.
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PMID:Activity of nimesulide on constitutive and inducible cyclooxygenases. 859 66


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