Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.24 (mitogen-activated protein kinase)
 95,810 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The growth rate of vascular smooth muscle cells (VSMCs), which were derived from aorta of mice deficient in the fibrinolytic factors tissue-type plasminogen activator (t-PA(-/-)), urokinase (u-PA(-/-)), u-PA receptor (u-PAR(-/-)) and type 1 plasminogen activator inhibitor (PAI-1(-/-)), as well as wild-type (WT) mice, was investigated in the presence of mouse melanoma cells (B16). In the VSMCs cultured with a basal medium supplemented with 10% fetal calf serum (FCS), there was no difference in the growth rate among the gene-lacking VSMCs and WT VSMCs, indicating that these fibrinolytic factors were not involved in the FCS-mediated cell proliferation. On the other hand, when these VSMCs were cultured with B16 cells in either the mixed culture or a double-chamber, only u-PAR(-/-) VSMCs showed a significantly lower growth rate. In addition, these suppressive effects on u-PAR(-/-) VSMCs were also observed in the presence of B16-derived conditioned medium (B16/CM). The growth rate of all the VSMCs except u-PAR(-/-) VSMCs was increased in the presence of B16/CM. The degree of the increase in cell number was comparable to that obtained with FCS. These effects on growth activity were partially associated with the levels of mitogen-activated protein kinase (MAPK, p42/p44) activity. The findings suggest that u-PAR plays an important role in the proliferative response of VSMCs and that without u-PAR, there is no intracellular signaling for cell proliferation.
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PMID:Growth inhibition of vascular smooth muscle cells derived from urokinase receptor (u-PAR)-deficient mice in the presence of carcinoma cells. 1508 64

Several proteases and their specific inhibitors modulate the interdependent processes of cell migration and matrix proteolysis as part of the global program of trauma repair. Expression of plasminogen activator inhibitor type-1 (PAI-1), a serine protease inhibitor (SERPIN) important in the control of barrier proteolysis and cell-to-matrix adhesion, for example, is spatially-temporally regulated following epithelial denudation injury in vitro as well as in vivo. PAI-1 mRNA/protein synthesis was induced early after epidermal monolayer scraping and restricted to keratinocytes comprising the motile cohort closely recapitulating, thereby, similar events during cutaneous healing. The time course of PAI-1 promoter-driven PAI-1-GFP fusion "reporter" expression in wound-juxtaposed cells approximated that of the endogenous PAI-1 gene confirming the location-specificity of gene regulation in this model. ERK activation was evident within 5 min after injury and particularly prominent in cells residing at the scrape-edge (suggesting a possible role in PAI-1 induction and/or the motile response) as was myosin light chain (MLC) phosphorylation. Indeed, MEK blockade with PD98059 or U0126 attenuated keratinocyte migration (by > or =60%), as did transient transfection of a dominant-negative ERK1 construct (40% decrease in monolayer repair), and completely inhibited PAI-1 transcript expression. Anti-sense down-regulation of PAI-1 synthesis (by 80-85%), or addition of PAI-1 neutralizing antibodies also inhibited injury site closure over a 24 h period establishing that PAI-1 was required for efficient long-term planar motility in this system. PAI-1 anti-sense transfection or actinomycin D transcriptional blockade, in contrast, did not affect the initial migratory response suggesting that residual PAI-1 protein levels (at least in transfectant cells and actinomycin D-treated cultures) may be sufficient to support early cell movement. Pharmacologic inhibition of keratinocyte MEK signaling effectively ablated scrape-induced PAI-1 mRNA expression but failed to attenuate wound-associated increases in cellular PAI-1 protein levels soon after monolayer injury. Collectively, these data suggest that basal PAI-1 transcripts may be mobilized for initial PAI-1 synthesis and, perhaps, the early motile response while maintenance of the normal rate of migration requires the prolonged PAI-1 expression that typically accompanies the repair response. To assess this possibility, scrape site closure studies were designed using keratinocytes isolated from PAI-1-/- mice. PAI-1-/- keratinocytes, in fact, had a significant wound healing defect evident even within the first 6 h following monolayer denudation injury. Addition of active PAI-1 protein to PAI-/- keratinocytes rescued the migratory phenotype that that approximating wild-type cells. These findings validate use of the present keratinocyte model to investigate injury-related controls on PAI-1 gene regulation and, collectively, implicate participation of PAI-1 in two distinct phases of epidermal wound repair.
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PMID:PAI-1 expression is required for epithelial cell migration in two distinct phases of in vitro wound repair. 1517

Obesity and insulin resistance confer increased risk for accelerated coronary disease and cardiomyopathic phenomena. We have previously shown that inhibition of angiotensin-converting enzyme (ACE) prevents coronary perimicrovascular fibrosis in genetically obese mice that develop insulin resistance. This study was performed to elucidate mechanism(s) implicated and to determine the effects of attenuation of angiotensin II (Ang) II. Genetically obese ob/ob mice were given ACE inhibitor (temocapril) or Ang II type 1 (AT(1)) receptor blocker (olmesartan) from 10 to 20 weeks. Cardiac expressions of plasminogen activator inhibitor (PAI)-1, the major physiologic inhibitor of fibrinolysis, and transforming growth factor (TGF)-beta(1), a prototypic profibrotic molecule, were determined and extent of perivascular coronary fibrosis was measured. Twenty-week-old obese mice exhibited increased plasma levels of PAI-1 and TGF-beta(1) compared with the values in lean counterpart. Perivascular coronary fibrosis in arterioles and small arteries was evident in obese mice that also showed increased left ventricular collagen as measured by hydroxyproline assay. Immunohistochemistry confirmed the deposition of perivascular type 1 collagen. Markedly increased PAI-1 and TGF-beta were seen immunohistochemically in coronary vascular wall and confirmed by western blotting. When obese mice were treated with temocapril or olmesartan from 10 to 20 weeks, both were equally effective and prevented increases in perivascular fibrosis, plasma PAI-1 and TGF-beta(1), left ventricular collagen and mural immunoreactivity for PAI-1, TGF-beta and collagen type 1. The c-Jun NH(2)-terminal kinase (JNK) activity was elevated in the left ventricle of obese mice (western) and blocked by temocapril and olmesartan. Ang II-mediated upregulation of PAI-1 and TGF-beta(1) with collagen deposition may explain the mechanism of perivascular fibrosis in obese mice. ACE inhibition and blockade of AT(1) receptor may prevent coronary perivascular fibrosis and collagen deposition even before development of overt diabetes. JNK activation may be a mediator of obesity-related cardiac dysfunction and a potential therapeutic target.
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PMID:Salutary effects of attenuation of angiotensin II on coronary perivascular fibrosis associated with insulin resistance and obesity. 1527 22

The urokinase receptor (uPAR) attenuates myofibroblast recruitment and fibrosis in the kidney. This study examined the role of uPAR and its co-receptor LDL receptor-related protein (LRP) in the regulation of kidney fibroblast proliferation and extracellular signal-regulated kinase (ERK) signaling. Compared with uPAR+/+ cells, uPAR-/- kidney fibroblasts were hyperproliferative. UPAR-/- fibroblast proliferation was 60% inhibited by an ERK kinase inhibitor. LRP protein was reduced and extracellular accumulation of urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor type 1 (PAI-1) proteins were greater in uPAR-/- cultures. Addition of functional uPA protein or LRP antisense RNA significantly increased ERK signaling and cell mitosis in both genotypes. Enhanced uPAR-/- fibroblast proliferation was reversed by a recombinant nonfunctional uPA peptide. The density of cell-bound fluor-uPA was similar between uPAR-/- and uPAR+/+ fibroblasts (78 +/- 6 versus 92 +/- 16 units). These data suggest that uPAR-deficient kidney fibroblasts express lower levels of its scavenger co-receptor LRP, resulting in greater extracellular accumulation of uPA and PAI-1. Enhanced proliferation of uPAR-/- fibroblasts seems to be mediated by uPA-dependent ERK signaling via an alternative urokinase receptor.
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PMID:Mitogenic signaling of urokinase receptor-deficient kidney fibroblasts: actions of an alternative urokinase receptor and LDL receptor-related protein. 1528 95

Although hepatocyte growth factor (HGF) can act synergistically or antagonistically with transforming growth factor-beta (TGF-beta) signaling, molecular mechanism of their crosstalk remains unknown. Using antibodies which selectively distinguished receptor-regulated Smads (R-Smads) phosphorylated at linker regions from those at C-terminal regions, we herein showed that either HGF or TGF-beta treatment of normal stomach-origin cells activated the JNK pathway, thereafter inducing endogenous R-Smads phosphorylation at linker regions. However, the phosphorylation at their C-terminal regions was not induced by HGF treatment. The activated JNK could directly phosphorylate R-Smads in vitro at the same sites that were phosphorylated in response to TGF-beta or HGF in vivo. Thus, the linker regions of R-Smads were the common phosphorylation sites for HGF and TGF-beta signaling pathways. The phosphorylation induced by simultaneous treatment with HGF and TGF-beta allowed R-Smads to associate with Smad4 and to translocate into the nucleus. JNK pathway involved HGF and TGF-beta-mediated infiltration potency since a JNK inhibitor SP600125 caused the reduction of invasive capacity induced by HGF and TGF-beta signals. Moreover, a combined treatment with HGF and TGF-beta led to a potent increase in plasminogen activator inhibitor type 1 transcriptional activity through Smad3 phosphorylation at the linker region. In contrast, HGF treatment reduced TGF-beta-dependent activation of p15INK4B promoter, in which Smad3 phosphorylation at the C-terminal region was involved. In conclusion, HGF and TGF-beta transmit the signals through JNK-mediated R-Smads phosphorylation at linker regions.
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PMID:TGF-beta and HGF transmit the signals through JNK-dependent Smad2/3 phosphorylation at the linker regions. 1532 85

The net balance between urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor type-1 (PAI-1) has been implicated in tumor cell invasion and metastasis. To elucidate the mechanism of the transforming growth factor-beta1 (TGF-beta1)-dependent up-regulation of PAI-1 expression, we investigated which signaling pathway transduced by TGF-beta1 is responsible for this effect. Here, we show (1) nontoxic concentrations of TGF-beta1 up-regulates uPA expression in HRA and SKOV-3 human ovarian cancer cells, (2) TGF-beta1 activates Smads (phosphorylation of Smad2 and nuclear translocation of Smad3) and subsequently up-regulates PAI-1 expression in HRA cells, whereas TGF-beta1 neither activates Smads nor up-regulates PAI-1 in SKOV-3 cells, (3) pharmacological Src inhibitor PP2 or antisense (AS) c-Src oligodeoxynucleotide (ODN) treatment significantly induces TGF-beta1-dependent activation of Smads, leading to PAI-1 synthesis, compared with controls, in SKOV-3 cells, (4) combination of TGF-beta1 and PP2, which activates PAI-1 expression and reduces uPA expression in SKOV-3, results in decreased invasiveness, (5) pharmacological inhibitors for mitogen-activated protein kinase (MAPK) (PD98059) and phosphoinositide-3-kinase (PI3K) (LY294002 and wortmannin) or AS-PI3K ODN transfection do not affect TGF-beta1-induced Smad signaling and up-regulation of PAI-1 expression in SKOV-3 cells pretreated with PP2, and (6) the induction of PAI-1 protein was partially inhibited by an inhibitor of Sp1-DNA binding, mithramycin, implicating, at least in part, Sp1 in the regulation of this gene by TGF-beta1. In conclusion, TGF-beta1-dependent activation of Smad2/3, leading to PAI-1 synthesis, may be negatively regulated by Src, but not its downstream targets MAPK and PI3K in SKOV-3 cells. These data also reflect the complex biological effect of uPA-PAI-1 system.
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PMID:Transforming growth factor-beta1-dependent activation of Smad2/3 and up-regulation of PAI-1 expression is negatively regulated by Src in SKOV-3 human ovarian cancer cells. 1537 29

The urokinase plasminogen activator(uPA) system plays important roles in tumor cell invasion and metastasis. In the present study, we evaluated the effects of ATF-PAI2CD, a hybrid protein of the amino-terminal fragment of urokinase and mutant plasminogen activator inhibitor-2, on 95D cells in vitro and in vivo. Furthermore, our results support a current hypothesis that fusion protein blocks tumor invasion and motility by inhibiting localized pericellular proteolysis. Treatment of 95D cells with ATF-PAI2CD resulted in a dose-dependent decrease in tumor-cell invasion through matrigel, and ATF-PAI2CD was much more effective than PAI-2CD. In addition, extracellular regular protein kinase (ERK1/2) expression was downregulated and the adhesion ability to fibronectin was increased in 95D cells treated with the fusion protein, which was confirmed by cell adhesion assay. A high-concentration of ATF-PAI2CD caused a significant reduction in tumor volume and weight in BALB/c (nu/nu) mice female inoculated with human 95D cells (5 x 10(6)); the antitumor effects were significant, which demonstrated a 67.9+/-4.2% reduction in tumor growth compared with control mice. The number of lymphatic metastasis was significantly reduced in mice treated with high- and middle- concentrations of ATF-PAI2CD, whereas a low-concentration of ATF-PAI2CD failed to exhibit any antimetastatic effects. In conclusion, the results suggested that the hybrid protein has therapeutic potential for lung carcinoma and other tumors to inhibit tumor invasion and metastasis.
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PMID:A hybrid protein of the amino-terminal fragment of urokinase and mutant plasminogen activator inhibitor-2 efficiently inhibits tumor cell invasion and metastasis. 1549 Feb 35

GnRH analog (GnRHa) and TGF-beta act directly on leiomyoma/myometrial smooth muscle cells (LSMCs and MSMCs) regulating diverse activities resulting in leiomyoma growth and regression. Because GnRH and TGF-beta receptor signaling is in part mediated through the MAPK pathway, we determined whether the contribution of MAPK/ERK and transcriptional activation of c-fos and c-jun, result in differential regulation of type I collagen, fibronectin, and plasminogen activator inhibitor 1 (PAI-1) gene expression, whose products are known to influence extracellular matrix turnover, which is critical in leiomyoma growth and GnRHa-induced regression. We found that GnRHa and TGF-beta in a dose- and time-dependent manner increased the level of phosphorylated ERK1/2 (pERK1/2) in LSMCs and MSMCs. GnRHa and TGF-beta increased ERK1/2 nuclear accumulation resulting in differential regulation of c-fos and c-jun mRNA expression via downstream signaling from MAPK kinase (MEK)1/2, because pretreatment with U0126, a synthetic inhibitor of MEK1/2, abolished basal and GnRHa- and TGF-beta-induced pERK1/2 and the expression of c-fos and c-jun. LSMCs and MSMCs also express fibronectin, type I collagen, and PAI-1 mRNA, and GnRHa and TGF-beta altered their expression in a cell-specific manner through MEK1/2. We concluded that GnRHa and TGF-beta acting through a MAPK/ERK pathway and transcriptional activation of c-fos/c-jun results in differential regulation of specific genes whose products may in part influence the outcome of leiomyoma growth and regression.
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PMID:Gonadotropin releasing hormone and transforming growth factor beta activate mitogen-activated protein kinase/extracellularly regulated kinase and differentially regulate fibronectin, type I collagen, and plasminogen activator inhibitor-1 expression in leiomyoma and myometrial smooth muscle cells. 1553 10

C-reactive protein (CRP) is significantly associated with the risk of ischemic cardiovascular disease in epidemiological studies. To explore if CRP has a functional role, we investigated its effect on the gene expression profile of vascular endothelial cells. Human vascular endothelial cells (human umbilical vein endothelial cells and human aortic endothelial cells) were incubated with CRP at various concentrations (0-10 mug/ml). Microarray analysis showed that a total of 11 genes increased (IL-8, core promoter element binding protein, activin A, monocyte chemoattractant protein 1, Exostoses 1, Cbp/p300-interacting transactivator with Glu/Asp-rich COOH-terminal domain 2, plasminogen activator inhibitor 1, fibronectin-1, gravin, connexin43, and sortilin-related receptor-1) and 6 genes decreased (methionine adenosyltransferase 2A, tryptophan-rich basic protein, reticulocalbin 1, membrane-associated RING-CH protein VI, cytoplasmic dynein1, and annexin A(1)) by more than twofold for their mRNA levels. IL-8 was the most significantly upregulated gene (13.6-fold), which demonstrated a clear dose- and time-dependent pattern revealed by quantitative real-time PCR. Cell adhesion assay showed that CRP enhanced the monocyte adhesion to endothelial cell monolayer by 2-fold (P < 0.01), which was partially blocked by an anti-IL-8 antibody (34.2% inhibition, P < 0.01). Inhibition of ERK MAPK pathway using U0126 prevented CRP-induced IL-8 upregulation, and Western blot analysis revealed a rapid activation of ERK1/2 after CRP stimulation. These data showed that CRP can significantly influence gene expressions in vascular endothelium. The CRP-responsive genes suggested that CRP may have a broad functional role in cell growth and differentiation, vascular remodeling and solid tumor development.
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PMID:Effect of C-reactive protein on gene expression in vascular endothelial cells. 1559 Oct 95

Transforming growth factor-beta1 (TGF-beta1) stimulates expression of plasminogen activator inhibitor type-1 (PAI-1), a serine protease inhibitor (SERPIN) important in the control of stromal barrier proteolysis and cell-to-matrix adhesion. Pharmacologic agents that target MEK (PD98059, U0126) or src family (PP1) kinases attenuated TGF-beta1-dependent PAI-1 transcription in R22 aortic smooth muscle cells. Pretreatment with PP1 at concentrations that inhibited TGF-beta1-dependent PAI-1 expression also blocked ERK1/2 activation/nuclear accumulation suggesting that the required src kinase activity is upstream of ERK1/2 in the TGF-beta1-initiated signaling cascade. The IC(50) of the PP1-sensitive kinase, furthermore, specifically implied involvement of pp60(c-src) in PAI-1 induction. Indeed, addition of TGF-beta1 to quiescent R22 cells resulted in a 3-fold increase in pp60(c-src) autophosphorylation and kinase activity. Transfection of a dominant-negative pp60(c-src) construct, moreover, reduced TGF-beta1-induced PAI-1 expression levels to that of unstimulated controls or PP1-pretreated cells. A >/=170 kDa protein that co-immunoprecipitated with TGF-beta1-activated pp60(c-src) was also phosphorylated transiently in response to TGF-beta1. TGF-beta1 is known to transactivate the 170 kDa EGF receptor (EGFR) by autocrine HB-EGF or TGF-alpha mechanisms suggesting involvement of EGFR activation in certain TGF-beta1-initiated responses. Incubation of quiescent R22 cells with the EGFR-specific inhibitor AG1478 prior to growth factor (EGF or TGF-beta1) addition effectively blocked EGFR activation as determined by direct visualization of receptor internalization. AG1478 suppressed (in a dose-dependent fashion) EGF-induced PAI-1 protein levels and, at a final concentration of 2.5 muM, virtually eliminated EGF-dependent PAI-1 synthesis. More importantly, AG1478 similarly repressed inducible PAI-1 levels in TGF-beta1-stimulated R22 cultures. PP1, PD98059, and U0126 also inhibited TGF-beta1-dependent cell motility at concentrations that significantly attenuated PAI-1 expression. Consistent with the AG1478-associated reductions in EGF- and TGF-beta1-stimulated PAI-1 expression, pretreatment of R22 cell cultures with AG1478 effectively suppressed growth factor-stimulated cell motility. These data indicate that two major phenotypic characteristics of TGF-beta1-exposure (i.e., transcription of specific target genes [e.g., PAI-1], increased cell motility) are linked in the R22 vascular smooth muscle cell system, require pp60(c-src) kinase activity and MEK signaling and involve activation of an AG1478-sensitive (likely EGFR-dependent) pathway.
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PMID:Plasminogen activator inhibitor type-1 gene expression and induced migration in TGF-beta1-stimulated smooth muscle cells is pp60(c-src)/MEK-dependent. 1562 20


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