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:2.7.10.1 (ERK)
95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Vascular endothelial growth factor (VEGF) is an important factor for endothelial cell proliferation and a key regulator of blood vessel development in embryos and angiogenesis in adult tissues. Its biological activity is mediated by two receptor tyrosine kinases, VEGFR-1 (Flt-1) and VEGFR-2 (KDR). In contrast to VEGFR-2, a naturally occurring soluble form of the VEGFR-1 (sVEGFR-1) is produced by endothelial cells by differential splicing of the flt-1 gene, and it is a secreted gene product. In order to develop a specific enzyme-linked immunosorbent assay (ELISA) for the measurement of sVEGFR-1, we established five anti-human receptor antibodies and characterized them in detail. These antibodies recognize different epitopes located within the seven Ig-like domains of the extracellular receptor protein but have no neutralizing activity in ligand binding assays. Together with a polyclonal antiserum, a specific human sVEGFR-1 ELISA was developed using the mAb #190.11. The ELISA can detect human sVEGFR-1 with a minimum detection limit of 1 ng/ml. The ELISA does not show any cross-reactivity with other related soluble receptors. Using this assay, human sVEGFR-1 was measured in the supernatant of different VEGFR-1 expressing cell types. No sVEGFR-1 protein was detectable after heparin Sepharose treatment or size-exclusion filtration (< 30 kDa). The ELISA assay for sVEGFR-1 was also used to measure the amount of the soluble receptor in amniotic fluid samples of patients undergoing amniocentesis during the course of normal pregnancies. The concentration of the samples was in the range of 5-35 ng/ml. This ELISA could be useful powerful tool for investigations concerning the physiological function of the soluble receptor under normal and pathophysiological conditions.Furthermore, it may facilitate studies of the mechanisms of receptor production.
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PMID:Detection and quantification of complexed and free soluble human vascular endothelial growth factor receptor-1 (sVEGFR-1) by ELISA. 1041 Sep 82

Vascular endothelial growth factor (VEGF) is an angiogenic peptide that can stimulate endothelial cell proliferation and migration in vitro and collateral development in ischemic organs in vivo. Although postulated, the expression of functional VEGF receptors in the heart has not been demonstrated yet. To prove this hypothesis and to extend the molecular basis of myocardial angiogenesis, we have characterized the expression and function of VEGF receptors in human coronary artery endothelial cells (HCAEC) and in human heart tissue. VEGF strongly induces proliferation and migration of HCAEC. These cells express transcripts of the two VEGF receptors KDR and Flt-1. Their expression levels are higher in HCAEC as compared with human umbilical vein endothelial cells. In HCAEC, VEGF stimulates phosphorylation of KDR in a concentration-dependent manner proving that KDDR is a functional receptor tyrosine kinase. Scatchard analysis demonstrated the presence of the high affinity receptor Flt-1 in HCAEC with a kd of 8 pM. Flt-1 protein could be visualized as a single band corresponding to a size of 210 kd. In addition mature KDR protein could be detected in adult human heart. Taken together, HCAEC and human heart tissue express high levels of functional VEGF receptors. These results broaden the molecular basis for understanding and manipulating VEGF-induced endothelial function and angiogenesis in the coronary circulation.
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PMID:The coronary endothelium: a target for vascular endothelial growth factor. Human coronary artery endothelial cells express functional receptors for vascular endothelial growth factor in vitro and in vivo. 1046 36

Vascular endothelial growth factor (VEGF) mediates increased vascular permeability and endothelial mitogenesis, and may orchestrate normal glomerular permselectivity and proteinuria. Distinct isoforms result from differential gene splicing. VEGF binds to two cell surface tyrosine-kinase receptors, KDR (kinase domain region) and Flt-1 (fms-like tyrosine kinase-1). The latter also exists in a soluble form (sFlt), which is inhibitory. We have studied patterns of VEGF-isoform and VEGF-receptor expression in isolated single normal human glomeruli. mRNA from 190 glomeruli (from 20 individuals) was harvested on to magnetic beads, and nested reverse transcription-PCR was performed using primers for the VEGF isoforms and VEGF receptors. Simultaneous nested reverse transcription-PCR for CD45 was conducted in order to exclude leucocyte contamination. Unexpected products were isolated, cloned and sequenced. Multiple patterns of glomerular VEGF mRNA isoform expression were identified. Most frequently (58%), all three common forms were expressed. VEGF(189) (i.e. 189-amino-acid form of VEGF) was expressed in 63%, VEGF(165) in 85% and VEGF(121) in 84% of glomeruli. Two unexpected PCR products were also identified: 18% of glomeruli expressed VEGF(145), and 27% of glomeruli expressed a new truncated VEGF splice variant, VEGF(148), lacking exon 6, the terminal part of exon 7 and exon 8. Multiple patterns of VEGF-receptor expression were also identified, the most common being expression of all three isoforms (28%). Overall, KDR was seen in 59% of glomeruli, Flt-1 in 45% and sFlt in 57%. Thus the expression of VEGF within normal glomeruli is complex and variable, with inter- and intra-individual variation. Furthermore, sFlt appears to be the co-dominant form of VEGF receptor expressed within glomeruli, suggesting that, in healthy individuals, a degree of VEGF autoregulation is the norm. The physiological importance of VEGF(148) remains to be confirmed.
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PMID:Heterogeneous vascular endothelial growth factor (VEGF) isoform mRNA and receptor mRNA expression in human glomeruli, and the identification of VEGF148 mRNA, a novel truncated splice variant. 1046 55

The angiogenic growth factor VEGF binds to the receptor tyrosine kinases Flt-1 and KDR/Flk-1. Immunoglobulin (Ig)-like loop-2 of Flt-1 is involved in binding VEGF, but the contribution of other Flt-1 Ig-loops to VEGF binding remains unclear. We tested the ability of membrane-bound chimeras between the extracellular domain of Flt-1 and the cell adhesion molecule embigin to bind VEGF. VEGF bound as well to receptors containing Flt-1 loops 1-2 or 2-3 as it did to the entire Flt-1 extracellular domain. Chimeras containing only loop-2 of Flt-1 bound VEGF with 22-fold lower affinity. We conclude that high-affinity VEGF binding requires Ig-like loop-2 plus either loop-1 or loop-3. In addition, Flt-1 amino acid residues Arg-224 and Asp-231 were not essential for high-affinity binding of VEGF to membrane-bound Flt-1.
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PMID:Characterization of the VEGF binding site on the Flt-1 receptor. 1047 94

Vascular endothelial growth factor A (VEGF) is a potent mitogen for endothelial cells in vitro and promotes neo-angiogenesis in vivo. VEGF overexpression occurs in most human malignancies including thyroid carcinomas in which elevated VEGF expression is associated with a high tumorigenic potential. To investigate the role of VEGF in angiogenesis associated with development of thyroid carcinomas, we constitutively expressed VEGF121 into a poorly tumorigenic cell line (NPA) expressing minimal levels of endogenous VEGF. Here we report that VEGF overexpressing NPA cells showed the same growth potential as untransfected NPA in vitro but formed well-vascularized tumors when injected subcutaneously into nude mice with markedly reduced latency compared to parental cells. A complementary approach was to suppress VEGF expression in a highly tumorigenic anaplastic cell line (ARO) by the transfection of an antisense construct. Antisense-transfected ARO cells expressed reduced constitutive levels of VEGF, showed the same growth potential as untransfected ARO cells in vitro and formed small tumors characterized by minimal vascularization, extensive necrosis and longer latency compared to parental or vector-transfected ARO cells in vivo. Finally, we investigated the expression of both VEGF tyrosine kinase receptors (Flt-1 and Flk-1/KDR) in tumor specimens by RT - PCR. Expression of (Flt-1 and Flk-1/KDR) was low in tissue specimens derived from NPA tumors, but was found enhanced in NPA VEGF tumors; conversely, the expression of VEGF receptors was high in tissue specimens derived from ARO tumors but was decreased in tumors derived from VEGF depleted ARO cells. These results clearly demonstrate that VEGF indirectly promotes the growth of thyroid tumors by stimulating angiogenesis.
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PMID:Modulation of in vivo growth of thyroid tumor-derived cell lines by sense and antisense vascular endothelial growth factor gene. 1049 Aug 19

Preeclampsia is a multisystem disorder characterized by hypertension and proteinuria. There is accumulating evidence that this is a disease of the endothelium, with an as-yet unidentified circulating factor, or factors, causing the observed alteration in vascular function. We previously reported that the function of myometrial vessels is altered on exposure to plasma from women with preeclampsia. Vascular endothelial growth factor (VEGF) is an angiogenic growth factor that acts via two high-affinity receptors (KDR and Flt-1), and its production is increased in preeclampsia. Here we report that VEGF and its Flt-1 receptor may play a pivotal role in the altered vascular function of preeclampsia. Myometrial resistance vessels were obtained at the time of cesarean section. Using the Mulvany wire myograph, the endothelium-dependent behavior of these vessels was studied. Incubation of vessels from pregnant women with VEGF resulted in a reduction of endothelium-dependent relaxation that mimicked the reduction induced by plasma from women with preeclampsia. The altered function that occurred upon exposure of vessels to VEGF or plasma from women with preeclampsia did not occur when plasma was incubated with antibodies to VEGF before vessel incubation. The presence of an anti-KDR receptor antibody had no effect on VEGF response. However, in the presence of an anti-Flt-1 receptor antibody, VEGF or plasma from women with preeclampsia no longer attenuated the endothelium-dependent relaxation (p < 0.05). The changes observed with VEGF and plasma from women with preeclampsia and their subsequent blockade with anti-VEGF antibody and anti-Flt-1 receptor antibody strongly suggest that VEGF acting through the Flt-1 receptor is pivotal in the pathogenesis of this disease.
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PMID:VEGF via VEGF receptor-1 (Flt-1) mimics preeclamptic plasma in inhibiting uterine blood vessel relaxation in pregnancy: implications in the pathogenesis of preeclampsia. 1049 28

Interaction of the hepatocyte growth factor (HGF) with its receptor, the Met tyrosine kinase, results in invasive growth, a genetic program essential to embryonic development and implicated in tumor metastasis. Met-mediated invasive growth requires autophosphorylation of the receptor on tyrosines located in the kinase activation loop (Tyr(1234)-Tyr(1235)) and in the carboxyl-terminal tail (Tyr(1349)-Tyr(1356)). We report that peptides derived from the Met receptor tail, but not from the activation loop, bind the receptor and inhibit the kinase activity in vitro. Cell delivery of the tail receptor peptide impairs HGF-dependent Met phosphorylation and downstream signaling. In normal and transformed epithelial cells, the tail receptor peptide inhibits HGF-mediated invasive growth, as measured by cell migration, invasiveness, and branched morphogenesis. The Met tail peptide inhibits the closely related Ron receptor but does not significantly affect the epidermal growth factor, platelet-derived growth factor, or vascular endothelial growth factor receptor activities. These experiments show that carboxyl-terminal sequences impair the catalytic properties of the Met receptor, thus suggesting that in the resting state the nonphosphorylated tail acts as an intramolecular modulator. Furthermore, they provide a strategy to selectively target the MET proto-oncogene by using small, cell-permeable, peptide derivatives.
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PMID:A peptide representing the carboxyl-terminal tail of the met receptor inhibits kinase activity and invasive growth. 1050 85

VEGF (vascular endothelial growth factor) overproduction has been identified as a major factor underlying pathological angiogenesis in vivo, including such conditions as psoriasis, macular degeneration, and tumor proliferation. Endothelial cell tyrosine kinase receptors, KDR and Flt-1, have been implicated in VEGF responses including cellular migration, proliferation, and modulation of vascular permeability. Therefore, agents that limit VEGF-cellular interaction are likely therapeutic candidates for VEGF-mediated disease states (particularly agents blocking activity of VEGF165, the most frequently occurring VEGF isoform). To that end, a nuclease-resistant, VEGF165-specific aptamer NX1838 (2'-fluoropyrimidine, RNA-based oligonucleotide/40-kDa-PEG) was developed. We have assessed NX1838 inhibition of a variety of cellular events associated with VEGF, including cellular binding, signal transduction, calcium mobilization, and induction of cellular proliferation. Our data indicate that NX1838 inhibits binding of VEGF to HUVECs (human umbilical vein endothelial cells) and dose-dependently prevents VEGF-mediated phosphorylation of KDR and PLCgamma, calcium flux, and ultimately VEGF-induced cell proliferation. NX1838-inhibition of VEGF-mediated cellular events was comparable to that observed with anti-VEGF monoclonal antibody, but was ineffective as an inhibitor of VEGF121-induced HUVEC proliferation. These findings, coupled with nuclease stability of the molecule, suggest that NX1838 may provide therapeutic utility in vivo.
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PMID:Oligonucleotide NX1838 inhibits VEGF165-mediated cellular responses in vitro. 1054 35

The mechanism by which vascular endothelial growth factor (VEGF) regulates endothelial nitric-oxide synthase (eNOS) expression is presently unclear. Here we report that VEGF treatment of bovine adrenal cortex endothelial cells resulted in a 5-fold increase in both eNOS protein and activity. Endothelial NOS expression was maximal following 2 days of constant VEGF exposure (500 pM) and declined to base-line levels by day 5. The elevated eNOS protein level was sustained over the time course if VEGF was co-incubated with L-N(G)-nitroarginine methyl ester, a competitive eNOS inhibitor. Addition of S-nitroso-N-acetylpenicillamine, a nitric oxide donor, prevented VEGF-induced eNOS up-regulation. These data suggest that nitric oxide participates in a negative feedback mechanism regulating eNOS expression. Various approaches were used to investigate the role of the two high affinity VEGF receptors in eNOS up-regulation. A KDR receptor-selective mutant increased eNOS expression, whereas an Flt-1 receptor-selective mutant did not. Furthermore, VEGF treatment increased eNOS expression in a KDR but not in an Flt-1 receptor-transfected porcine aorta endothelial cell line. SU1498, a selective inhibitor of the KDR receptor tyrosine kinase, blocked eNOS up-regulation, thus providing further evidence that the KDR receptor signals for eNOS up-regulation. Finally, treatment of adrenal cortex endothelial cells with VEGF or phorbol ester resulted in protein kinase C activation and elevated eNOS expression, whereas inhibition of protein kinase C with isoform-specific inhibitors abolished VEGF-induced eNOS up-regulation. Taken together, these data demonstrate that VEGF increases eNOS expression via activation of the KDR receptor tyrosine kinase and a downstream protein kinase C signaling pathway.
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PMID:Vascular endothelial growth factor governs endothelial nitric-oxide synthase expression via a KDR/Flk-1 receptor and a protein kinase C signaling pathway. 1055 75

VEGF is mitogenic, angiogenic, and a potent mediator of vascular permeability. VEGF causes extravasation of plasma protein in skin bioassays and increases hydraulic conductivity in isolated perfused microvessels. Reduced tissue oxygen tension triggers VEGF expression, and increased protein and mRNA levels for VEGF and its receptors (Flt-1, Flk-1/KDR) occur in the ischemic rat brain. Brain edema, provoked in part by enhanced cerebrovascular permeability, is a major complication in central nervous system pathologies, including head trauma and stroke. The role of VEGF in this pathology has remained elusive because of the lack of a suitable experimental antagonist. We used a novel fusion protein, mFlt(1-3)-IgG, which sequesters murine VEGF, to treat mice exposed to transient cortical ischemia followed by reperfusion. Using high-resolution magnetic resonance imaging, we found a significant reduction in volume of the edematous tissue 1 day after onset of ischemia in mice that received mFlt(1-3)-IgG. 8-12 weeks after treatment, measurements of the resultant infarct size revealed a significant sparing of cortical tissue. Regional cerebral blood flow was unaffected by the administration of mFlt(1-3)-IgG. These results demonstrate that antagonism of VEGF reduces ischemia/reperfusion-related brain edema and injury, implicating VEGF in the pathogenesis of stroke and related disorders.
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PMID:VEGF antagonism reduces edema formation and tissue damage after ischemia/reperfusion injury in the mouse brain. 1058 25


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