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)

The non-angiogenic role of vascular endothelial growth factor (VEGF), and its receptors flt-1 and flk-1, together with downstream signaling pathways were examined in fetal and postnatal rat cerebral cortical organotypic explants. VEGF application in both paradigms caused a significant increase in astroglial proliferation and a dose-dependent increase in GFAP and nestin immunoreactivity. The VEGF receptor flt-1 was observed on most, though not all astrocytes, while flk-1 receptor immunoexpression was absent. Treatment with antisense oligonucleotides (AS-ODNs) to flt-1 resulted in a dramatic decrease in GFAP and nestin immunoreactivity, which further confirmed the role of flt-1 in mediating VEGF's gliotrophic effects, while AS-ODNs to flk-1 had no effect. VEGF-induced gliotrophic effects were found to be mediated by the MAPK/ERK and PI-3 kinase signaling pathways, since the both the MEK1 inhibitor, PD98059 and the PI-3 kinase inhibitor, Wortmannin abolished VEGF-induced astrocytic GFAP(+) expression. Although high dose VEGF application resulted in strong upregulation of both GFAP and nestin immunoreactivity in astrocytes, overlap of the two proteins was not observed in all cells, suggesting that some of the nestin(+) cells might be neural progenitors. Exposure to VEGF resulted in upregulation of both VEGF and bFGF mRNA at the one-day time point, and bFGF protein by 3 days; VEGF activated astrocytes expressed bFGF to a much greater degree than those in untreated explants. The increased expression of bFGF induced by VEGF, may serve in the proliferation of multipotential neural stem/progenitor cells in vitro. VEGF, an established angiogenic factor, appears to play a significant role in the growth and differentiation of astrocytes in the CNS.
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PMID:Astrocyte growth effects of vascular endothelial growth factor (VEGF) application to perinatal neocortical explants: receptor mediation and signal transduction pathways. 1575 57

Bupleuran 2IIc, a pectic polysaccharide isolated from the roots of bupleurum falcatum L., was previously characterized as a T-cell-independent B cell mitogen. This study focuses on elucidating the mechanism by which bupleuran 2IIc induces cyclin D2 production for inducing mitogenesis in murine B cells. Bupleuran 2IIc was digested with endo-alpha-(1-->4)-D-polygalacturonase and the resulting bupleuran 2IIc/PG-1 ("ramified" region) strongly stimulated cyclin D2 expression. When murine B cells were stimulated with bupleuran 2IIc/PG-1, phosphorylation of tyrosine residues of a number of proteins was observed. Cyclin D2 expression by bupleuran 2IIc/PG-1 was inhibited by the tyrosine kinase inhibitors, genistein and herbimycin A, and the Src family tyrosine kinase inhibitor, PP2, suggesting a possible role for tyrosine kinases. The stimulation by bupleuran 2IIc/PG-1 of cyclin D2 expression was significantly decreased by inhibitors, PI 3-kinase (LY294002 and Wortmannin), PLCgamma (U73122), PKC (H-7), receptor-operated calcium entry inhibitor (SK&F 96365), and calcineurin (FK506). Both PD98059 and U0126, highly selective inhibitors of MEK1 and MEK1/2, respectively, did not strongly suppress the expression of cyclin D2 after stimulation by bupleuran 2IIc/PG-1. The results suggest that (1) bupleuran 2IIc/PG-1 is the active site for induction of cyclin D2 by bupleuran 2IIc, (2) the expression of the cyclin D2 gene by bupleuran 2IIc/PG-1 may be mediated via the activation of PI 3-kinase and PLCgamma followed by activation of PKC and calcium mobilization, and (3) the ERK1/2 cascade is not a central signaling pathway for bupleuran 2IIc/PG-1-induced cyclin D2 expression.
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PMID:A possible signal transduction pathway for cyclin D2 expression by a pectic polysaccharide from the roots of bupleurum falcatum L. in murine B cell. 1595 64

The function of muscarinic acetylcholine receptors expressed in oligodendrocytes and in myelin has remained largely undetermined. Here we present evidence that incubation of oligodendrocyte progenitors, deprived of growth factor, with the acetylcholine analog carbachol significantly reduced cell death by apoptosis and blocked caspase-3 cleavage. This protective effect was reversed by atropine, a muscarinic acetylcholine receptor antagonist, as well as by specific inhibitors of intracellular signaling molecules, including phosphatidylinositol 3-kinase (Wortmannin and LY294002), Akt (Akt inhibitor III) and Src-like tyrosine kinases (PP2), but not by the mitogen-activated protein kinase kinase inhibitor, PD98059. Activation of Akt by carbachol was antagonized by atropine and inhibited by LY294002 and PP2. The Src-like tyrosine kinase inhibitor, PP2, also reduced carbachol stimulation of extracellular signal-regulated kinases 1/2 and cAMP-response element binding protein in a dose-dependent manner. Furthermore, carbachol increased tyrosine-phosphorylation of Fyn, a member of the Src-like tyrosine kinases. These results indicate that muscarinic acetylcholine receptors play an important role in oligodendrocyte progenitor survival through transduction pathways involving activation of Src-like tyrosine kinases and phosphatidylinositol 3-kinase/Akt.
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PMID:Muscarinic acetylcholine receptors mediate oligodendrocyte progenitor survival through Src-like tyrosine kinases and PI3K/Akt pathways. 1643 36

During pregnancy, vascular remodeling and vasoactive agents such as nitric oxide (NO) increase blood flow to the uteroplacental unit. Using our uterine artery endothelial cell (UAEC) culture model, based on cells from pregnant (P-UAEC) and nonpregnant (NP-UAEC) ewes, we investigate the relative physiological roles of Ca(2+) vs. kinase in the regulation of endothelial NO synthase (eNOS) activity. When Ca(2+) mobilization is fully inhibited using inhibitors of phospholipase C (PLC) (U73122) and the inositol triphosphate (IP3) receptor (IP3-R) (2-APB), significant residual eNOS activity remains in both P- and NP-UAEC. No change in ATP-stimulated ERK2, Akt, or eNOS phosphorylation is observed with U73122 (0.01-1 microM) or 2-APB (1-50 microM). The MAPK kinase (MEK) 1/2 inhibitor U0126 (10 microM) did not alter ATP-stimulated eNOS activity in P-UAEC, but potentiated the ATP response in NP-UAEC. Using two phosphatidylinositol 3-kinase (PI3-K) inhibitors, we observed no effect with LY294002 (10 microM) on eNOS activity in P- and NP-UAEC, but wortmannin (10 microM) inhibited both P- and NP-UAEC eNOS activation. Expression of constitutively active Akt (ca-Akt) in UAEC resulted in slight elevation of basal eNOS activity, but relative ATP-stimulated eNOS activation was not altered by ca-Akt. Wortmannin continued to inhibit eNOS activation by ATP in the presence of ca-Akt; LY294002 still had no inhibitory effect. Our data indicate both [Ca(2+)](i) and multiple kinases are involved in the regulation of eNOS activity in our model. We report that pregnancy adaptation of eNOS activation includes the reduced sensitivity to ERK-mediated attenuation of eNOS activity and enhanced stimulation of eNOS activity through a wortmannin-sensitive, LY294002-insensitive, Akt-independent mechanism.
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PMID:Pregnancy-enhanced endothelial nitric oxide synthase (eNOS) activation in uterine artery endothelial cells shows altered sensitivity to Ca2+, U0126, and wortmannin but not LY294002--evidence that pregnancy adaptation of eNOS activation occurs at multiple levels of cell signaling. 1645 84

Mechanical stretching represents an important part of the signaling in skin. We have previously demonstrated that mechanical stretching induced proliferative phenotypes in human keratinocytes, as shown in increased 5-bromo-2'-deoxyuridine (BrdU) incorporation, ERK1/2 activation, and keratin K6 induction. Here we have further investigated the antiapoptotic signals in human keratinocytes provoked by mechanical stretching in vitro. Keratinocytes were plated on flexible silicone supports to transmit mechanical stretching to keratinocytes, involving continuous stretching by +20%. Stretching of these cells for 15-30 min caused the phosphorylation and activation of Akt. Inhibition of mitogen and extracellular signal-regulated kinase (MEK1/2) with U0126 and phosphoinositide 3-OH kinase (PI 3-K) with Wortmannin attenuated Akt activation. The epidermal growth factor (EGF) receptor kinase inhibitor, AG1478, and calcium channel inhibitor, gadolinium (Gd3+), also inhibited Akt activation. In summary, our results demonstrate the activation of the Akt signaling pathway by mechanical stretching, generating not only proliferative but also antiapoptotic signals in human keratinocytes.
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PMID:Activation of Akt by mechanical stretching in human epidermal keratinocytes. 1663 75

Obesity has been recognized as a risk factor for breast cancer. Adipocyte-derived leptin may play as a paracrine regulator on the growth of breast cancer cells. Expression of both leptin and its OB-Rb receptor was detected in human breast cancer ZR-75-1 cells and further induced by leptin, suggesting that both expression and message mediation of leptin were autoregulated by itself. With cell counting and MTT assay, we had observed leptin stimulated ZR-75-1 growth in dose- and time-dependent manners. To study what steps of cell cycle progression leptin may involve in, we analyzed cell-cycle profile with flow cytometric analysis, mRNA and protein expressions of four cell-cycle regulators with RT-PCR and Western blotting analysis. Under the treatment of leptin, the G1 arrest of cells was reduced accompanied with up-regulation of G1 phase-specific cyclin D1 and proto-oncogene c-Myc, but down-regulation of cyclin-dependent kinase inhibitor p21(WAF1/CIP1) and tumor suppressor p53. Furthermore, JAK2 inhibitor AG490, PI3K/Akt inhibitor Wortmannin, and MEK/ERK1/2 inhibitor PD98059 were efficiently prevented leptin-promoted cell growth. Effect of cooperation between leptin and estrogen on ZR-75-1 growth had been observed. Collectively, the results showed that the proliferative effect of leptin on ZR-75-1 was associated with the up-regulation of cyclin D1 and c-Myc and down-regulation of tumor suppressor p53 and p21(WAF1/CIP1) plausibly through a hypothesized JAK2-PI3K/Akt-MEK/ERK pathway. The leptin- and OB-Rb-expressing capability of ZR-75-1 created a possible autocrine control of leptin, in which signal could be effectively amplified by itself, on cell growth.
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PMID:Leptin-induced growth of human ZR-75-1 breast cancer cells is associated with up-regulation of cyclin D1 and c-Myc and down-regulation of tumor suppressor p53 and p21WAF1/CIP1. 1675 79

RAW macrophages, which express the PDE4D3 and PDE4D5 cAMP phosphodiesterase isoforms, exhibited increased PDE4 activity when challenged with H2O2 in a fashion that was negated by treatment with the cell permeant antioxidant, N-acetyl cysteine and by diphenyleneiodonium chloride, an inhibitor of NADPH oxidase. In Cos1 cells transfected to express PDE4D3, challenge with H2O2 caused a rapid increase in both the activity and phosphorylation of PDE4D3. Lysates from H2O2-treated COS cells caused the phosphorylation of purified, recombinant PDE4D3 at two sites. One was the established ERK phosphorylation site at Ser579, located at the extreme C-terminus of the catalytic unit, and the other was a novel site at Ser239, located at the extreme N-terminus of the catalytic unit. Double Ser239Ala:Ser579Ala mutation of PDE4D3 prevented its H2O2-dependent phosphorylation both in vitro and in intact COS cells. Phosphorylation of PDE4D3 at Ser579 was ablated by treating COS cells with the MEK inhibitor, PD98059, which also negated activation. The activity of the Ser239Ala:Ser579Ala double mutant, and the Ser579Ala single PDE4D3 mutant was unaffected by H2O2 challenge of COS cells, whilst the Ser239Ala mutant was inhibited. Wortmannin inhibited the H2O2-dependent phosphorylation of PDE4D3 in COS cells by around 50%, whilst it fully ablated phosphorylation at Ser239 as well as ablating activation of PDE4D3. Neither immunodepletion of p70S6 kinase nor siRNA-mediated knockdown of mTor inhibited the H2O2-dependent phosphorylation of PDE4D3 at Ser239. Activation of PDE4D3 by challenge with H2O2 was not additive with activation through protein kinase A (PKA)-mediated phosphorylation of PDE4D3. Challenge with H2O2 did not alter PKA-mediated phosphorylation of PDE4D3 at Ser54. H2O2 dependent phosphorylation of PDE4D3, at Ser239 and Ser579, did not alter the sensitivity of PDE4D3 to inhibition by the selective PDE4 inhibitor, rolipram. An unknown protein kinase acting downstream of phosphatidyl inositol 3-kinase phosphorylates PDE4D3 at Ser239. This switches the effect of phosphorylation by ERK at Ser579 from inhibition to activation. We propose that phosphorylation at Ser239 attenuates interaction between either UCR2 or the UCR1/UCR2 module and the PDE4 catalytic unit so as to re-programme the functional outcome effect of phosphorylation by ERK. We identify a novel process through which reactive oxygen species activate long PDE4 isoforms so as to reduce cAMP levels and thereby promote inflammatory responses.
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PMID:Oxidative stress employs phosphatidyl inositol 3-kinase and ERK signalling pathways to activate cAMP phosphodiesterase-4D3 (PDE4D3) through multi-site phosphorylation at Ser239 and Ser579. 1697 30

We previously showed that tumor necrosis factor-alpha (TNF-alpha) stimulates synthesis of interleukin-6 (IL-6), a potent bone resorptive agent, via p44/p42 mitogen-activated protein (MAP) kinase in osteoblast-like MC3T3-E1 cells. In the present study, we investigated whether phosphatidylinositol 3-kinase (PI3-kinase)/protein kinase B (Akt) is involved in TNF-alpha-stimulated IL-6 synthesis in MC3T3-E1 cells. TNF-alpha induced the phosphorylation of Akt depending upon time. Akt inhibitor, 1L-6-hydroxymethyl-CHIRO-inositol 2-( R)-2- O-methyl-3-O-octadecylcarbonate, significantly suppressed the TNF-alpha-stimulated IL-6 synthesis, but the inhibitory effect was partial. The phosphorylation of Akt induced by TNF-alpha was markedly attenuated by LY294002 and wortmannin, inhibitors of PI3-kinase. Wortmannin and LY294002 significantly reduce the TNF-alpha-induced IL-6 synthesis. On the contrary, the suppressive effects of Akt inhibitor, wortmannin or LY294002 on TNF-alpha-induced phosphorylation of p44/p42 MAP kinase were minor. PD98059, a specific inhibitor of MEK, had little effect on the TNF-alpha-induced phosphorylation of Akt. A combination of Akt inhibitor and PD98059 suppressed the TNF-alpha-induced IL-6 synthesis in an additive manner. These results strongly suggest that PI3-kinase/Akt plays a role in the TNF-alpha-stimulated IL-6 synthesis mainly independent of p44/p42 MAP kinase in osteoblasts.
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PMID:Phosphatidylinositol 3-kinase/Akt plays a part in tumor necrosis factor-alpha-induced interleukin-6 synthesis in osteoblasts. 1698 Nov 37

Little is known about the regulatory mechanisms of endothelial cell (EC) proliferation by retinal pericytes and vice versa. In a model of coculture with bovine retinal pericytes lasting for 24 h, rat brain ECs showed an increase in arachidonic acid (AA) release, whereas Western blot and RT-PCR analyses revealed that ECs activated the protein expression of cytosolic phospholipase A(2) (cPLA(2)) and its phosphorylated form and calcium-independent intracellular phospholipase A(2) (iPLA(2)). No activation of the same enzymes was seen in companion pericytes. In ECs, the protein level of phosphorylated extracellular signal-regulated kinase (ERK) 1/2 was also enhanced significantly, a finding not observed in cocultured pericytes. The expression of protein kinase C-alpha (PKCalpha) and its phosphorylated form was also enhanced in ECs. Wortmannin, LY294002, and PD98059, used as inhibitors of upstream kinases (the PI3-kinase/Akt/PDK1 or MEK-1 pathway) in cultures, markedly attenuated AA release and the expression of phosphorylated forms of endothelial cPLA(2), PKCalpha, and ERK1/2. By confocal microscopy, activation of PKCalpha in perinuclear regions of ECs grown in coculture as well as strong activation of cPLA(2) in ECs taken from a model of mixed culture were clearly observed. However, no increased expression of both enzymes was found in cocultured pericytes. Our findings indicate that a sequential activation of PKCalpha contributes to endothelial ERK1/2 and cPLA(2) phosphorylation induced by either soluble factors or direct cell-to-cell contact, and that the PKCalpha-cPLA(2) pathway appears to play a key role in the early phase of EC-pericyte interactions regulating blood retina or blood-brain barrier maturation.
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PMID:Endothelial cell-pericyte cocultures induce PLA2 protein expression through activation of PKCalpha and the MAPK/ERK cascade. 1726 47

Insulin-like growth factor-I (IGF-I) is required for the growth of oligodendrocytes, although the underlying mechanisms are not fully understood. Our aim was to investigate the role of phosphatidylinositol 3-kinase (PI3K), mitogen-activated protein kinase kinase (MEK1), and Src family tyrosine kinases in IGF-I-stimulated proliferation of oligodendrocyte progenitors. IGF-I treatment increased the proliferation of cultured oligodendrocyte progenitors as determined by measuring incorporation of [(3)H]-thymidine and bromodeoxy-uridine (BrdU). IGF-I stimulated a transient phosphorylation of 3-phosphoinositide-dependent kinase-1 (PDK1) and extracellular signal-regulated kinases (ERK1/2) (targets of MEK1), as well as a rapid and sustained activation of Akt (a target of PI3K). Furthermore, inhibitors of PI3K (LY294002 and Wortmannin), MEK1 (PD98059 and U0126), and Src family tyrosine kinases (PP2) decreased IGF-I-induced proliferation, and blocked ERK1/2 activation. LY294002, Wortmannin and PP2 also blocked Akt activation. To further determine whether Akt is required for IGF-I stimulated oligodendrocyte progenitor proliferation, cultures were infected with adenovirus vectors expressing dominant-negative mutants of Akt or treated with pharmacological inhibitors of Akt. All treatments reduced IGF-I-induced oligodendrocyte progenitor proliferation. Our data indicate that stimulation of oligodendrocyte progenitor proliferation by IGF-I requires Src-like tyrosine kinases as well as the PI3K/Akt and MEK1/ERK signaling pathways.
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PMID:IGF-I-induced oligodendrocyte progenitor proliferation requires PI3K/Akt, MEK/ERK, and Src-like tyrosine kinases. 1734 61


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