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: EC:2.7.12.2 (MEK)
18,161 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Treatment of mouse astrocyte cultures with combined interleukin (IL)-1alpha and tumor necrosis factor (TNF)-alpha induced expression of inducible nitric-oxide synthase (iNOS), resulting in sustained release of large amounts of nitric oxide, whereas TNF-alpha and IL-1alpha individually were unable to induce iNOS expression in astrocytes. The role of MAPK cascades and of NF-kappaB activation in the early intracellular signal transduction involved in iNOS transcription in TNF-alpha/IL-1alpha-stimulated astrocytes was investigated. TNF-alpha and IL-1alpha activated all p42/44(MAPK), p38(MAPK), and p54(JNK) pathways as determined by immunoprecipitation kinase assays using specific antibodies and substrates. The p38(MAPK) pathway is specifically involved in TNF-alpha/IL-1alpha-induced iNOS expression, since iNOS protein and nitric oxide release in the presence of a specific inhibitor of p38(MAPK), 4-(4-fluorophenyl)-2-2-(4-hydroxyphenyl)-5-(4-pyridyl)-imidazole (FHPI), were dramatically diminished. In contrast, PD98059, a specific inhibitor of MEK1 had no effect on iNOS expression. p38(MAPK) did not couple NF-kappaB to iNOS transcription, but NF-kappaB had a clear role in iNOS transcription regulation. Northern blot analysis showed that the p38(MAPK) pathway controlled iNOS expression at the transcriptional level, since iNOS mRNA was reduced in the presence of FHPI in TNF-alpha/IL-1alpha-stimulated astrocytes. iNOS expression was investigated with TNF receptor (TNFR)-1- and TNFR-2-deficient mice. The TNF-alpha activity in TNF-alpha/IL-1alpha-stimulated astrocytes was exclusively mediated through TNFR-1, most likely because TNFR-2-mediated signals in astrocytes did not connect to the p38(MAPK) pathway. These data suggest that TNF-alpha/IL-1alpha-induced iNOS expression depends on a yet undetermined second pathway in addition to p38(MAPK).
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PMID:Blockade of p38 mitogen-activated protein kinase pathway inhibits inducible nitric-oxide synthase expression in mouse astrocytes. 935 95

The sphingomyelin signal transduction pathway is known to play a role in mediating the action of various cytokines. Here we examined the possible role of the sphingomyelin signaling pathway on lipopolysaccharide (LPS)- and cytokine-mediated production of NO and the expression of inducible nitric-oxide synthase (iNOS). Sphingomyelinase (SMase) treatment of astrocytes increased the cellular levels of ceramide without the induction of NO production. However, incubation of LPS or cytokine-stimulated astrocytes with SMase or by increasing intracellular ceramide by cell-permeable ceramide analogs (C2- or C6-ceramide) or inhibitor of ceramidase (N-oleoyl ethanolamine) led to a time- and dose-dependent increase in the production of NO. This increase in NO production was accompanied by an increase in iNOS activity, iNOS protein, and iNOS mRNA. Similar to astrocytes, SMase or ceramide analogs also stimulated the LPS- and cytokine-mediated expression of iNOS in the C6 glial cell line. Since activation of NF-kappaB is necessary for the induction of iNOS, we examined the effect of SMase and C2-ceramide on the activation of NF-kappaB. Although SMase or C2-ceramide alone was ineffective in activating NF-kappaB, both stimulated the LPS-mediated activation of NF-kappaB in LPS-activated astrocytes. Inhibition of ceramide and LPS-mediated induction of iNOS by antioxidant inhibitors of NF-kappaB (N-acetylcysteine and pyrrolidine dithiocarbamate) suggest that the stimulatory effect of ceramide on the induction of iNOS is due to the stimulation of NF-kappaB activation and that cellular redox plays a role in the activation of NF-kappaB and induction of iNOS. Inhibition of LPS-mediated as well as LPS and ceramide-mediated induction of iNOS and activation of NF-kappaB by PD98059, a specific inhibitor of activation of mitogen-activated protein (MAP) kinase kinase (MEK), and FPT inhibitor II, a selective inhibitor of Ras farnesyl protein transferase, indicate that the Ras-MAP kinase pathway is involved in LPS-ceramide induced activation of NF-kappaB and induction of iNOS, and that ceramide-mediated signaling events probably converge into the LPS-modulated MAP kinase signaling pathway resulting in greater activation of NF-kappaB and iNOS induction. This study illustrates a novel role of the sphingomyelin-ceramide signaling pathway in stimulating the expression of iNOS via LPS- or cytokine-mediated activation of NF-kappaB in astrocytes.
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PMID:Sphingomyelinase and ceramide stimulate the expression of inducible nitric-oxide synthase in rat primary astrocytes. 944 61

The influence of osmolarity and compatible organic osmolytes on the phosphorylation of the MAP-kinases Erk-1 and Erk-2 and on the expression of taurine transporter (TAUT) and lipopolysaccharide (LPS)-induced nitric oxide synthetase (iNOS) was studied in RAW 264.7 mouse macrophages. Hypoosmolarity (205 mosmol/l) but not hyperosmolarity (405 mosmol/l) or challenge of the cells with betaine or taurine increased phosphorylation of Erk-1 and Erk-2. Hypoosmotic Erk-phosphorylation was blocked by the MEK-inhibitor PD098059 but was resistant to depletion of extracellular calcium and to inhibition of PLC, PKC, erbstatin-sensitive tyrosine kinases and elevation of intracellular cAMP. Hyperosmolarity stimulated Na+-dependent taurine uptake and led to an increase of TAUT mRNA levels, whereas hypoosmotic exposure diminished both and induced a rapid efflux of the osmolyte from taurine-preloaded cells. The hyperosmotic elevation of TAUT mRNA levels was antagonized upon addition of taurine but not of betaine or myo-inositol. Hyperosmolarity increased the LPS-induced iNOS expression at the mRNA and the protein level. This was suppressed by betaine but not by taurine or myo-inositol. The osmotic regulation of taurine transport and iNOS expression appeared independent of the MEK-Erk pathway and the p38MAPK.
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PMID:Compatible organic osmolytes and osmotic modulation of inducible nitric oxide synthetase in RAW 264.7 mouse macrophages. 970 50

This study examined the signal transduction pathway(s) leading to phosphorylation of p38 in human neutrophils stimulated with lipopolysaccharide and formyl peptides. Blockade of the nitric oxide (NO) pathway in neutrophils with the NO synthase inhibitor N-nitro-L-arginine methyl ester or by treatment with the NO scavenger 2-phenyl-tetramethylimidazoline-1-oxyl-3-oxide attenuated phosphorylation of the mitogen-activated protein kinase p38 in response to lipopolysaccharide but not fMet-Leu-Phe. Using the NO releasing agents S-nitroso-N-acetylpenicillamine and sodium nitroprusside it was determined that nitric oxide is sufficient to cause an increase in phosphorylation of p38. Increasing cellular cGMP with phosphodiesterase inhibitors, by stimulation of soluble guanylyl cyclase with YC-1 or with exogenous dibutyryl cGMP resulted in mitogen-activated protein kinase/extracellular signal-regulated kinase kinase 3,6 (MEK3,6) activation and phosphorylation of p38. This phenomenon was specific for MEK3,6, because these agents had no effect on the phosphorylation state of MEK1,2. A role for protein kinase G but not protein kinase A downstream of lipopolysaccharide but not formylmethionylleucylphenylalanine was shown using the specific inhibitors KT5823 and H89, respectively. These data indicate that activation of p38 by fMet-Leu-Phe and lipopolysaccharide involve different mechanisms, and that activation of protein kinase G by NO-dependent stimulation of guanylyl cyclase is necessary and sufficient for phosphorylation of p38 downstream of lipopolysaccharide.
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PMID:Activation of p38 mitogen-activated protein kinase by lipopolysaccharide in human neutrophils requires nitric oxide-dependent cGMP accumulation. 986 77

The effect of norepinephrine (NE) on cytokine-stimulated nitric oxide (NO) production by cardiac myocytes has not been previously reported. NE alone caused no significant increase in NO-2 levels over vehicle. Addition of NE to interleukin-1beta (IL-1beta) significantly increased inducible NO synthase (iNOS) mRNA expression, iNOS protein, and NO-2 production vs. IL-1beta alone. Addition of the alpha-adrenergic blocker prazosin or the beta-adrenergic blocker propranolol partially reduced the NE-mediated increase in iNOS mRNA expression and NO-2 production. Addition of prazosin and propranolol together completely abolished the NE-induced increase in iNOS mRNA expression and NO-2 production. NE significantly enhanced mitogen-activated protein (MAP) kinase activity that was reduced by prazosin, propranolol, and PD-98059, a selective MAP kinase kinase inhibitor. Addition of PD-98059 reduced the NE-mediated increase in iNOS mRNA expression and NO-2 production. We report for the first time that NE enhances IL-1beta-stimulated NO production by activation of alpha- and beta-adrenergic receptors through a novel MAP kinase mechanism.
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PMID:Norepinephrine-stimulated MAP kinase activity enhances cytokine-induced NO production by rat cardiac myocytes. 988 16

Angiotensin II (ATII) and platelet-derived growth factor (PDGF) are two vasoconstrictors implicated in the maintenance of normal vascular homeostasis. PDGF A-chain levels increase in cultured vascular smooth muscle cells (SMCs) exposed to ATII. The molecular mechanisms underlying this induction are not known. We used transient transfection analysis to show that ATII can increase reporter gene activity driven by fragments of the PDGF-A promoter bearing recognition elements for the transcription factor, Egr-1. Nuclear run-off experiments indicate that ATII induces Egr-1 expression at the level of transcription. Gel shift and supershift studies show that Egr-1 protein accumulates in the nuclei of SMCs exposed to ATII and binds to the proximal region of the PDGF-A promoter in a specific, time-dependent manner. ATII induced extracellular-signal regulated kinase (p42/44 ERK) activity as did phorbol 12-myristate 13-acetate. The specific MEK1/2 inhibitor, PD98059, suppressed both PDGF-A and Egr-1 endogenous and promoter-dependent expression inducible by ATII. The ATII type 1 receptor (AT1) antagonist, Losartan, inhibited ATII-induction of p42/44 ERK, as well as Egr-1 and PDGF-A, whereas neither PD123319, an AT2 receptor antagonist, nor wortmannin, an inhibitor of phosphatidylinositol 3-kinase and c-Jun N-terminal kinase, had any effect. ATII-induction of Egr-1 and PDGF-A was blocked by SIN-1, a NO donor. In addition, this pathway was blocked by overexpression of NO synthase. Collectively, these findings demonstrate that ATII activation of the PDGF-A promoter is mediated via the MEK/ERK/Egr-1 pathway and AT1 receptor and that this process is antagonized by NO.
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PMID:Angiotensin II (ATII)-inducible platelet-derived growth factor A-chain gene expression is p42/44 extracellular signal-regulated kinase-1/2 and Egr-1-dependent and mediated via the ATII type 1 but not type 2 receptor. Induction by ATII antagonized by nitric oxide. 1044 31

Interleukin 1beta (IL-1beta) induces expression of the inducible nitric-oxide synthase (iNOS) with concomitant release of nitric oxide (NO) from glomerular mesangial cells. These events are preceded by activation of the c-Jun NH(2)-terminal kinase/stress-activated protein kinase (JNK/SAPK) and p38(MAPK). Our current study demonstrates that overexpression of the dominant negative form of JNK1 or p54 SAPKbeta/JNK2 significantly reduces the iNOS protein expression and NO production induced by IL-1beta. Similarly, overexpression of the kinase-dead mutant form of p38alpha(MAPK) also inhibits IL-1beta-induced iNOS expression and NO production. In previous studies we demonstrated that IL-1beta can activate MKK4/SEK1, MKK3, and MKK6 in renal mesangial cells; therefore, we examined the role of these MAPK kinases in the modulation of iNOS induced by IL-1beta. Overexpression of the dominant negative form of MKK4/SEK1 decreases IL-1beta-induced iNOS expression and NO production with inhibition of both SAPK/JNK and p38(MAPK) phosphorylation. Overexpression of the kinase-dead mutant form of MKK3 or MKK6 demonstrated that either of these two mutant kinase inhibited IL-1beta-induced p38(MAPK) (but not JNK/SAPK) phosphorylation and iNOS expression. Interestingly overexpression of wild type MKK3/6 was associated with phosphorylation of p38(MAPK); however, in the absence of IL-1beta, iNOS expression was not enhanced. This study suggests that the activation of both SAPK/JNK and p38alpha(MAPK) signaling cascades are necessary for the IL-1beta-induced expression of iNOS and production of NO in renal mesangial cells.
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PMID:Both p38alpha(MAPK) and JNK/SAPK pathways are important for induction of nitric-oxide synthase by interleukin-1beta in rat glomerular mesangial cells. 1059 6

Our laboratory has recently demonstrated a role for the phosphatidylinositol 3-kinase-mediated inducible NO synthase (iNOS) signaling pathway in acute regulation of insulin-induced mitogen-activated protein phosphatase-1 (MKP-1) expression in primary cultures of rat aortic vascular smooth muscle cells (VSMCs) (N. Begum, L. Ragolia, M. McCarthy, and N. Duddy. J. Biol. Chem. 273: 25164-25170, 1998). We now show that prolonged treatment of VSMCs with 100 nM insulin and high glucose (25 mM) for 12-24 h, to mimic hyperinsulinemia and hyperglycemia, completely blocked MKP-1 mRNA and protein expression in response to subsequent acute insulin treatment. To understand the mechanism of insulin resistance induced by high glucose and insulin, we studied the regulation of iNOS protein induction in these cells. Both high glucose and chronic insulin treatment caused a marked impairment of iNOS induction in response to acute insulin. Blocking of signaling via the p38 mitogen-activated protein kinase (MAPK) pathway by prior treatment for 1 h with SB-203580, a synthetic p38 MAPK inhibitor, completely prevented the inhibition of iNOS induced by high glucose and insulin and restored MKP-1 induction to levels observed with acute insulin treatment. In contrast, PD-98059, a MEK inhibitor, had no effect. Furthermore, high glucose and chronic insulin treatment caused sustained p38 MAPK activation. We conclude 1) that chronic insulin and high glucose-induced insulin resistance is accompanied by marked reductions in both iNOS and MKP-1 inductions due to p38 MAPK activation that leads to excessive cell growth and 2) that p38 MAPK/extracellular signal-regulated kinase pathways regulate iNOS induction, thereby controlling MKP-1 expression, which in turn inactivates MAPKs as a feedback mechanism and inhibits cell growth.
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PMID:High glucose and insulin inhibit VSMC MKP-1 expression by blocking iNOS via p38 MAPK activation. 1064 15

Macrophages produce large amounts of nitric oxide (NO) in response to proinflammatory cytokines and lipopolysaccharide (LPS) by expressing inducible isoform of NO synthase (iNOS). We examined the role of extracellular signal-regulated kinase p42/44(MAPK) (Erk1/2) in signal transduction pathways leading to induction of NO synthesis in response to LPS in J774 mouse macrophages and T-84 human colon epithelial cells. LPS activated Erk1/2 and induced iNOS and subsequent NO production. Erk1/2 activation was inhibited by PD 98059, a specific inhibitor of mitogen-activated protein kinase kinase (Mek) that is an upstream activator of Erk1/2. At corresponding concentrations PD 98059 reduced LPS-induced NO formation by 40 to 50% by inhibiting iNOS expression in J774 and T-84 cells. Inhibition of iNOS expression was not mediated by nuclear factor-kappaB because PD 98059 had no effect on nuclear factor-kappaB activity in J774 macrophages. In addition, PD 98059 reduced LPS-induced L-arginine transport into the cells as measured in J774 macrophages, whereas the availability of tetrahydrobiopterin was not a limiting factor in NO production after PD 98059. Our results indicate that Erk1/2 activation mediates up-regulation but is not essential for LPS-induced iNOS expression.
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PMID:Inhibition of extracellular signal-regulated kinase suppresses endotoxin-induced nitric oxide synthesis in mouse macrophages and in human colon epithelial cells. 1094 76

The 29-kDa amino-terminal fibronectin fragment (FN-f) has a potent chondrolytic effect and is thought to be involved in cartilage degradation in arthritis. However, little is known about signal transduction pathways that are activated by FN-f. Here we demonstrated that FN-f induced nitric oxide (NO) production from human articular chondrocytes. Expression of inducible nitric-oxide synthase (iNOS) mRNA and NO production were observed at 6 and 48 h after FN-f treatment, respectively. Interleukin-1beta (IL-1beta) mRNA up-regulation was stimulated by FN-f in human chondrocytes. To address the possibility that FN-f-induced NO release is mediated by IL-1beta production, the effect of IL-1 receptor antagonist (IL-1ra) was determined. IL-1ra partially inhibited FN-f-induced NO release although it almost completely inhibited IL-1beta-induced NO release. Tyrosine phosphorylation of focal adhesion kinase was induced transiently by FN-f treatment. Blocking antibodies to alpha(5) or beta(1) integrin and Arg-Gly-Asp-containing peptides did not inhibit FN-f-induced NO production. PP2, a Src family kinase inhibitor, or cytochalasin D, which selectively disrupts the network of actin filaments, inhibited both FAK phosphorylation and NO production induced by FN-f, but the phosphatidylinositol 3-kinase inhibitor wortmannin had no effect. Analysis of mitogen-activated protein kinases (MAPK) showed activation of extracellular signal-regulated kinase (ERK), c-Jun NH(2)-terminal kinase, and p38 MAPK. High concentrations of SB203580, which inhibit both JNK and p38 MAPK, and PD98059 a selective inhibitor of MEK1/2 that blocks ERK activation, inhibited FN-f induced NO production. These data suggest that focal adhesion kinase and MAPK mediate FN-f induced activation of human articular chondrocytes.
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PMID:Focal adhesion kinase and mitogen-activated protein kinases are involved in chondrocyte activation by the 29-kDa amino-terminal fibronectin fragment. 1167 48


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