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

This study was conducted to compare the levels of prostaglandin E2 (PGE2) released by cultured granulosa cells collected from normally-ovulating women (normal cells, NC) and those with polycystic ovaries (polycystic ovary granulosa cells, POGC). Granulosa cells were collected from 7 normal women and 7 anovulatory women with polycystic ovaries. Both groups underwent laparoscopic oocyte retrieval for gamete intra-fallopian transfer. Cell cultures were carried out under basal conditions and in the presence of various substances known to influence PGE2 biosynthesis. Prostaglandin E2 concentrations in the incubation media were taken as a marker of cyclo-oxygenase activity. Unexpectedly, POGC appeared to release greater amounts of PGE2 compared to the NC. There was no difference between the levels of PGE2 produced by the two types of cells during the first 3 hours after cell explants, whereas a difference (P < 0.01) was observed after 24 and 48 hours of incubation. Interleukin-1 beta enhanced PGE2 secretion (P < 0.01) in both POGC and NC, while lipopolysaccharide increased prostaglandin release only by the NC cells. Indomethacin inhibited PGE2 production to a greater extent in POGC (from -70 to -90% with respect to basal release, P < 0.01) than NC (approximately -50%, P < 0.01). Blockade by indomethacin and the weak inhibitory effect of the glucocorticoid, dexamethasone (P < 0.05 only in NC, and only at 24 hours), provided pharmacological evidence that PG production by granulosa cells in vitro might depend primarily on constitutive cyclo-oxygenase activity.
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PMID:Increased production and release of prostaglandin-E2 by human granulosa cells from polycystic ovaries. 890 19

Lipopolysaccharide treated rats (25 mg/kg i.v.) were killed after 60 min and rat aortic rings were mounted in an isolated organ bath for measurement of isometric contractions in response to phenylephrine (0.01-10 microM) or potassium chloride (10 mM). Aortic rings from lipopolysaccharide-treated rats showed reduced contractility to phenylephrine and potassium chloride when compared to those from saline-treated rats. Indomethacin 10 microM, added in vitro further impaired phenylephrine-induced contraction of aortic rings from lipopolysaccharide ex vivo treated rats but was ineffective on aortic rings from saline treated rats. A similar pattern was observed when potassium chloride was used. Administration in vitro of thromboxane A2 receptor antagonist SQ29,548 gave a similar effect to indomethacin. Aortic rings collected from rat treated in vivo with dexamethasone (10 mg/kg) showed a reduction in phenylephrine induced contractions that was not further reduced by in vitro treatment with indomethacin (10 microM). Similarly, when rat aortic rings were incubated in vitro (60 min) with lipopolysaccharide (0.4 mg/ml) a reduction of phenylephrine- and potassium chloride-induced contraction was observed, but addition of either indomethacin or SQ29,548 did not further reduce contraction. Our results suggest that under these experimental conditions, in the early phase of endotoxin shock, synthesis of cyclooxygenase products (such as endoperoxides or thromboxane A2) occurs probably as a compensatory mechanism to lipopolysaccharide induced hypocontractility from the interaction, in vivo, between lipopolysaccharide, endothelium, circulating cells and vascular smooth muscles.
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PMID:Indomethacin and thromboxane A2/prostaglandin H2 antagonist SQ29,548 impair in vitro contractions of aortic rings of ex vivo-treated lipopolysaccharide rats. 892 98

The involvement of prostaglandins in feedback modulation of the lipopolysaccharide-inducible isoforms of cyclooxygenase and nitric oxide was studied. This was done by testing the effects of arachidonic acid and indomethacin in the murine macrophage cell line J774. When added before lipopolysaccharide, arachidonic acid (10(-6)-10(-4) M) dose-dependently reduced inducible nitric oxide synthase and cyclooxygenase-2 activity as well as reducing the expression of both enzyme proteins. Indomethacin (10(-7)-10(-4) M) suppressed prostaglandin E2 release into the medium and reduced cyclooxygenase-2 activity irreversibly, as expected, but the amount of cyclooxygenase-2 protein was increased (because indomethacin removes the inhibitory effect of prostaglandins on cyclooxygenase-2 expression). However, the same doses of indomethacin also inhibited the release of nitric oxide into the medium. This was accompanied by a reduction in the amount of inducible nitric oxide synthase protein. The drug was not cytotoxic. Thus these results show that pharmacological treatments can have opposing actions on inducible nitric oxide synthase and cyclooxygenase-2 induction, with indomethacin showing an additional unexpected action in macrophages which may diminish their cytotoxic potential.
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PMID:Induction of cyclooxygenase and nitric oxide synthase in endotoxin-activated J774 macrophages is differentially regulated by indomethacin: enhanced cyclooxygenase-2 protein expression but reduction of inducible nitric oxide synthase. 898 31

1. It has been proposed that in inflammatory conditions, in which both the inducible isoforms of nitric oxide synthase (iNOS) and cyclo-oxygenase (COX-2) are induced, inhibition of NOS also results in inhibition of arachidonic acid metabolism. In the present study we have investigated whether mercaptoalkylguanidines, a novel class of selective iNOS inhibitors, may also influence the activity of cyclo-oxygenase (COX). Therefore, the effect of mercaptoethylguanidine (MEG) and related compounds on the activity of the constitutive (COX-1) and the inducible COX (COX-2) was investigated in cells and in purified enzymes. Aminoguanidine, NG-methyl-L-arginine (L-NMA) and NG-nitro-L-arginine methyl ester (L-NAME) were also studied for comparative purposes. 2. Western blot analysis demonstrated a significant COX-1 activity in unstimulated J774 macrophages and in unstimulated human umbilical vein endothelial cells (HUVEC). Immunostimulation of the J774 macrophages by endotoxin (lipopolysaccharide of E. coli, LPS 10 micrograms ml-1) and interferon gamma (IFN gamma, 100 u ml-1) for 6 h resulted in a significant induction of COX-2, and a down-regulation of COX-1. No COX-2 immunoreactivity was detected in unstimulated HUVEC or unstimulated J774 cells. Therefore, in subsequent studies, the effect of mercaptoalkylguanidines on COX-1 activity was studied in HUVEC stimulated with arachidonic acid for 6 h, and in J774 cells stimulated with arachidonic acid for 30 min. The effect of mercaptoalkylguanidines on COX-2 activity was studied in immunostimulated J774 macrophages, both on prostaglandin production by endogenous sources, and on prostaglandin production in response to exogenous arachidonic acid stimulation. In addition, the effect of mercaptoalkylguanidines on purified COX-1 and COX-2 activities was also studied. 3. In experiments designed to measure COX-1 activity in HUVEC, the cells were stimulated by arachidonic acid (15 microM) for 6 h. This treatment induced a significant production of 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha, the stable metabolite of prostacyclin), while nitrite production was undetectable by the Griess reaction. MEG (1 microM to 3 mM) caused a dose-dependent inhibition of the accumulation of 6-keto-PGF1 alpha, with an IC50 of 20 microM. However, aminoguanidine, L-NAME or L-NMA (up to 3 mM) did not affect the production of 6-keto-PGF1 alpha in this experimental system. In experiments designed to measure COX-1 activity in J774.2 macrophages, the cells were stimulated by arachidonic acid (15 microM) for 30 min; this also induced a significant production of 6-keto-PGF1 alpha and MEG (1 microM to 3 mM), aminoguanidine (at 1 and 3 mM), but neither L-NAME nor L-NMA inhibited the production of prostaglandins. 4. In experiments designed to measure prostaglandin production by COX-2 with endogenous arachidonic acid, J774.2 cells were immunostimulated for 6 h in the absence or presence of various inhibitors. In experiments designed to measure prostaglandin production by COX-2 with exogenous arachidonic acid, J774.2 cells were immunostimulated for 6 h, followed by a replacement of the culture medium with fresh medium containing arachidonic acid and various inhibitors. Both of these treatments induced a significant production of 6-keto-PGF1 alpha. Nitrite production, an indicator of NOS activity, was moderately increased after immunostimulation. MEG (1 microM to 3 mM) caused a dose-dependent inhibition of the accumulation of COX metabolites. Similar inhibition of LPS-stimulated 6-keto PGF1 alpha production was shown by other mercaptoalkylguanidines (such as N-methyl-mercaptoethylguanidine, N,N'-dimethyl-mercaptoethylguanidine, S-methyl-mercaptoethylguanidine and guanidino-ethyldisulphide), with IC50 values ranging between 34-55 microM. However, aminoguanidine, L-NAME and L-NMA (up to 3 mM) did not affect the production of prostaglandins.5. In comparative experiments indomethacin, a non selective COX inhibitor, and NS-398, a selective COX-2 inhibitor, reduced (LPS) stimulated 6-keto-PGF1alpha production in J774 macrophages in a dose-dependent manner without affecting nitrite release. Indomethacin, but not NS-398, inhibited 6-keto-PGF1alpha production in the HUVECs. 6.The inhibitory effect of MEG was due to direct inhibition of the catalytic activity of COX as indicated in experiments with purified COX-1 and COX-2. MEG dose-dependently inhibited the purified COX-1 and COX-2 activity with IC50 values of 33microM and 36microM, respectively. Aminoguanidine (at the highest concentrations) inhibited the formation of COX-1 metabolites, without affecting COX-2 activity. High doses of L-NAME (3mM) decreased COX-1 activity only, while L-NMA (up to 3mM) had no effect on the activity of either enzyme. 7.These results suggest that MEG and related compounds are direct inhibitors of the constitutive and the inducible cyclo-oxygenases, in addition to their effects on the inducible NOS. The additional effect of mercaptoalkylguanidines on COX activity may contribute to the beneficial effects of these agents in inflammatory conditions where both iNOS and COX-2 are expressed.
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PMID:The inhibitory effects of mercaptoalkylguanidines on cyclo-oxygenase activity. 903 36

We tested the hypothesis that increased dietary fish oil levels (via modulation of the production of inflammatory mediators) modulate sickness symptoms (i.e., anorexia, cachexia, fever, lethargy) of systemic and local inflammation. Swiss Webster mice were implanted with biotelemeters to measure body temperature and motor activity and were fed a diet high in n-3 fatty acids (17% wt/wt menhaden oil) or a reference diet (17% wt/wt hydrogenated coconut oil or normal rodent chow) for 6 wk. Local inflammation was induced by subcutaneous injection of turpentine (100 microl/mouse). Systemic inflammation was elicited by intraperitoneal injection of lipopolysaccharide (LPS; 2.5 mg/kg). Fever, lethargy, anorexia, and weight decrease during turpentine abscess were all inhibited (P < 0.05) in mice fed the fish oil diet. Indomethacin, similar to the fish oil diet, attenuated the turpentine-induced symptoms in mice fed a normal diet. Dietary n-3 fatty acids prevented fever and attenuated the decrease in body weight caused by LPS but did not affect the LPS-induced lethargy and anorexia. Within 90 min of LPS injection, the bioactivity of plasma tumor necrosis factor-alpha (TNF-alpha) increased to 98.2 +/- 5.1 ng/ml in mice fed fish oil compared with 32.6 +/- 3.6 ng/ml in those fed the reference diet (P < 0.05). Plasma prostaglandin E2 (PGE2) levels after LPS injection of mice fed the control diet increased within 90 min to 16.4 +/- 5.1 pg/ml. Mice fed the fish oil diet did not show any elevation in plasma PGE2 levels at that time (P < 0.05). We speculate that dietary n-3 fatty acids suppressed PGE2-related responses, including a PGE2-dependent negative feedback on TNF-alpha production, which resulted in differential modulation of sickness behavior depending on the locus of inflammation.
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PMID:Dietary n-3 fatty acids differentially affect sickness behavior in mice during local and systemic inflammation. 914 33

1. DFU (5,5-dimethyl-3-(3-fluorophenyl)-4-(4-methylsulphonyl)phenyl-2(5H)-furan one) was identified as a novel orally active and highly selective cyclo-oxygenase-2 (COX-2) inhibitor. 2. In CHO cells stably transfected with human COX isozymes, DFU inhibited the arachidonic acid-dependent production of prostaglandin E2 (PGE2) with at least a 1,000 fold selectivity for COX-2 (IC50 = 41 +/- 14 nM) over COX-1 (IC50 > 50 microM). Indomethacin was a potent inhibitor of both COX-1 (IC50 = 18 +/- 3 nM) and COX-2 (IC50 = 26 +/- 6 nM) under the same assay conditions. The large increase in selectivity of DFU over indomethacin was also observed in COX-1 mediated production of thromboxane B2 (TXB2) by Ca2+ ionophore-challenged human platelets (IC50 > 50 microM and 4.1 +/- 1.7 nM, respectively). 3. DFU caused a time-dependent inhibition of purified recombinant human COX-2 with a Ki, value of 140 +/- 68 microM for the initial reversible binding to enzyme and a kappa 2 value of 0.11 +/- 0.06 s-1 for the first order rate constant for formation of a tightly bound enzyme-inhibitor complex. Comparable values of 62 +/- 26 microM and 0.06 +/- 0.01 s-1, respectively, were obtained for indomethacin. The enzyme-inhibitor complex was found to have a 1:1 stoichiometry and to dissociate only very slowly (t1/2 = 1-3 h) with recovery of intact inhibitor and active enzyme. The time-dependent inhibition by DFU was decreased by co-incubation with arachidonic acid under non-turnover conditions, consistent with reversible competitive inhibition at the COX active site. 4. Inhibition of purified recombinant human COX-1 by DFU was very weak and observed only at low concentrations of substrate (IC50 = 63 +/- 5 microM at 0.1 microM arachidonic acid). In contrast to COX-2, inhibition was time-independent and rapidly reversible. These data are consistent with a reversible competitive inhibition of COX-1. 5. DFU inhibited lipopolysaccharide (LPS)-induced PGE2 production (COX-2) in a human whole blood assay with a potency (IC50 = 0.28 +/- 0.04 microM) similar to indomethacin (IC50 = 0.68 +/- 0.17 microM). In contrast, DFU was at least 500 times less potent (IC50 > 97 microM) than indomethacin at inhibiting coagulation-induced TXB2 production (COX-1) (IC50 = 0.19 +/- 0.02 microM). 6. In a sensitive assay with U937 cell microsomes at a low arachidonic acid concentration (0.1 microM), DFU inhibited COX-1 with an IC50 value of 13 +/- 2 microM as compared to 20 +/- 1 nM for indomethacin. CGP 28238, etodolac and SC-58125 were about 10 times more potent inhibitors of COX-1 than DFU. The order of potency of various inhibitors was diclofenac > indomethacin approximately naproxen > nimesulide approximately meloxicam approximately piroxicam > NS-398 approximately SC-57666 > SC-58125 > CGP 28238 approximately etodolac > L-745,337 > DFU. 7. DFU inhibited dose-dependently both the carrageenan-induced rat paw oedema (ED50 of 1.1 mg kg-1 vs 2.0 mg kg-1 for indomethacin) and hyperalgesia (ED50 of 0.95 mg kg-1 vs 1.5 mg kg-1 for indomethacin). The compound was also effective at reversing LPS-induced pyrexia in rats (ED50 = 0.76 mg kg-1 vs 1.1 mg kg-1 for indomethacin). 8. In a sensitive model in which 51Cr faecal excretion was used to assess the integrity of the gastrointestinal tract in rats, no significant effect was detected after oral administration of DFU (100 mg kg-1, b.i.d.) for 5 days, whereas chromium leakage was observed with lower doses of diclofenac (3 mg kg-1), meloxicam (3 mg kg-1) or etodolac (10-30 mg kg-1). A 5 day administration of DFU in squirrel monkeys (100 mg kg-1) did not affect chromium leakage in contrast to diclofenac (1 mg kg-1) or naproxen (5 mg kg-1). 9. The results indicate that COX-1 inhibitory effects can be detected for all selective COX-2 inhibitors tested by use of a sensitive assay at low substrate concentration. The novel inhibitor DFU shows the lowest inhibitory potency against COX-1, a consistent high selectivity of inhibition of COX-2 over COX-1 (>300 fold) with enzyme, whole cell and whole blood assays, with no detectable loss of integrity of the gastrointestinal tract at doses >200 fold higher than efficacious doses in models of inflammation, pyresis and hyperalgesia. These results provide further evidence that prostanoids derived from COX-1 activity are not important in acute inflammatory responses and that a high therapeutic index of anti-inflammatory effect to gastropathy can be achieved with a selective COX-2 inhibitor.
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PMID:Biochemical and pharmacological profile of a tetrasubstituted furanone as a highly selective COX-2 inhibitor. 914 94

The expression of surface procoagulants by exudative macrophages represents an important mechanism underlying local fibrin deposition at sites of extravascular inflammation. The present studies investigated the contribution of tyrosine phosphorylation to the generation of macrophage procoagulant activity (PCA) and tissue factor expression in response to proinflammatory stimuli. Both lipopolysaccharide (LPS) and zymosan rapidly stimulated tyrosine phosphorylation in elicited murine peritoneal macrophages. This effect was prevented by the tyrosine kinase inhibitors genistein and herbimycin and augmented by the addition of the phosphotyrosine phosphatase inhibitor vanadate. The vanadate-mediated rise in phosphotyrosine accumulation was abrogated by the use of diphenylene iodonium, an inhibitor of the respiratory burst oxidase, suggesting a role for peroxides of vanadate as contributors to the tyrosine phosphorylation. This notion was supported by the finding that vanadyl hydroperoxide markedly increased the accumulation of phosphotyrosine residues. To define the role of tyrosine phosphorylation in the induction of macrophage PCA by LPS, the effects of tyrosine kinase inhibition by genistein and herbimycin were investigated. Both agents inhibited the expression of macrophage PCA. Further, Northern blot analysis with the cDNA probe for murine tissue factor indicated that the inhibition occurred at the mRNA level or earlier. Since vanadate augmented phosphotyrosine accumulation, it was hypothesized that it might enhance generation of macrophage products. However, vanadate reduced induction of PCA in response to LPS. By contrast, vanadate augmented basal prostaglandin E2 (PGE2) release and stimulated PGE2 release by macrophages. Indomethacin prevented the increase in PGE2 but only partially restored normal levels of PCA. The effect of vanadate on tissue factor expression appeared to be posttranscriptional. These studies thus demonstrate, by functional Western blotting and Northern blotting techniques, that tyrosine phosphorylation plays a role in the regulation of macrophage PCA and tissue factor expression in response to proinflammatory stimuli.
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PMID:The role of tyrosine phosphorylation in lipopolysaccharide- and zymosan-induced procoagulant activity and tissue factor expression in macrophages. 916 75

Intussusception is a major cause for intestinal obstruction in children. Its etiology is unclear, but it is often associated with some kind of infection. We have developed a model for intussusception in mice using intraperitoneal (IP) injection of lipopolysaccharide (LPS). The objective of this study was to identify the putative mediators that participate in this LPS-induced intussusception. LPS (12 mg/kg) was injected into adult mice (N = 52) and 6 hr later, 25% of the animals demonstrated intussusception in the small or large intestine. We next tested whether nitric oxide (NO) or various inflammatory mediators contributed to this effect: Indomethacin (10 mg/kg) injected with LPS (12 mg/kg) completely prevented the effect of LPS (N = 20). The tumor necrosis factor (TNF) blocker pentoxifylline (200 mg/kg) significantly reduced the incidence of intussusception to 6.6% (N = 30). The platelet-activating factor (PAF) antagonist BN52021 (10 and 20 mg/kg) reduced the incidence of intussusception to 13.3% in both doses (N = 15 for each dose). Addition of 2% arginine (NO precursor) to the drinking water 36 hr before the injection of LPS increased the incidence of intussusception to 30.7% (N = 32). In mice injected with the NO synthase inhibitor L-NAME (20 mg/kg) only 3.8% developed intussusception (N = 26). Our results indicate that the induction of intussusception by LPS proceeds via parallel pathways involving cytokines, prostaglandins, and NO. Our previous pathological study showed that LPS did not cause any changes that may act as a lead point for the intussusception, suggesting that LPS induced intussusception by altering gut motility. We therefore propose that these mediators combine to induce disturbed gut motility that results in the formation of intussusception.
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PMID:The contribution of inflammatory mediators and nitric oxide to lipopolysaccharide-induced intussusception in mice. 920 71

Portal venous transfusions (PVTs) of blood have been shown to induce significant immunosuppression in animal models of organ transplantation. A proposed mechanism of PVT-induced immunosuppression is via alteration of Kupffer cell arachidonic acid metabolism with increased secretion of the suppressive metabolite prostaglandin E2 (PGE2). This study assessed the hypothesis that PVT increases Kupffer cell PGE2 production via up-regulation of Kupffer cell phospholipase A2 (PLA2) as well as constitutive (COX1) and inducible (COX2) cyclooxygenase. Kupffer cells from Lewis rats were harvested 1 hr after PVT with either 1 ml of Wistar-Furth blood, systemic transfusion (SVT), or saline via portal vein (PVSal). After lipopolysaccharide stimulation, 24-hr Kupffer cell supernatant fractions were assayed for PGE2. PGE2 was increased after SVT (1465+/-234 pg/ml) compared with PVSal (597+/-99; P<0.01). PVT increased Kupffer cell PGE2 (5370+/-533; P<0.001 vs. SVT and vs. PVSal) even more substantially. Kupffer cells from PVT-treated rats were then cultured in the presence of inhibitors of PLA2, COX1, or COX2. When Kupffer cells were treated with mepacrine to inhibit PLA2 (5575+/-453 pg/ml), PGE2 production was not different from that by PVSal-treated controls (6467+/-614 pg/ml), but when Kupffer cells were incubated in the presence of the COX1 inhibitor flurbiprofen (3512+/-407 pg/ml) or the COX2 inhibitor nimesulide (2800+/-830 pg/ml), production was decreased 46.7% and 56.7%, respectively, over control activity without added inhibitor. PVT also increased Kupffer cells COX1 and COX2 mRNA as measured by Northern blot. Heart transplants were then performed from Wistar-Furth donors into Lewis recipients at the time of PVT, SVT, PVSal, or PVT + indomethacin (COX1/2 inhibitor). PVT prolonged allograft survival (12.0+/-0.9 days) compared with PVSal (6.3+/-0.3; P<0.01) or SVT (6.3+/-0.3; P<0.04). Indomethacin shortened graft survival when given with PVT (6.5+/-0.3 days). In summary, PVT increased Kupffer cell PGE2 production, up-regulated transcription of Kupffer cells COX1 and COX2 mRNA, and prolonged cardiac allograft survival. COX1/2 inhibition abrogated the effect of PVT. The results indicated that the immunosuppressive effect of PVT may be mediated by up-regulation of Kupffer cell COX1 and COX2. Manipulation of Kupffer cell arachidonic acid metabolism may be useful in augmentation of PVT-induced immunosuppression.
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PMID:Portal venous transfusion up-regulates Kupffer cell cyclooxygenase activity: a mechanism of immunosuppression in organ transplantation. 923 13

1. The growing pig provides a useful, nonrodent model for studying mechanisms involved in the febrile response. 2. Indomethacin (IND) has previously been shown to prevent lipopolysaccharide (LPS) fever in prepubertal pigs. 3. This study compared the abilities of IND and L-745,337, a novel cyclooxygenase-2 (cox-2) inhibitor, to counteract the effects a low dose of LPS (20 micrograms/pig IV) on deep body temperature. Effects of IND and L-745,337 on core temperature and plasma cortisol concentrations were also examined in nonfebrile animals. 4. L-745,337 (0.3 mg/kg IV) did not alter the response to LPS, whereas both IND and L-745,337 (1.7 mg/kg) reduced the febrile effects of LPS given 60 min earlier. 5. Neither IND, nor L-745,337 (1.7 mg/kg IV) affected core temperature in nonfebrile animals whereas IND, but not L-745,337, stimulated cortisol release. 6. The results suggest that prostaglandin modulates the febrile effects of LPS in swine and that inhibition of inducible cyclooxygenase (Cox-2) suppresses fever without producing the stressful side-effects that accompany constitutive cyclooxygenase (Cox-1) inhibition, as exemplified by IND (a mixed Cox-1/Cox-2 antagonist).
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PMID:Comparison of the antipyretic actions of indomethacin and L-745,337, a selective cyclooxygenase-2 inhibitor, in endotoxin-treated prepubertal pigs. 945 83


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