Early Growth Response-1 Regulates Angiopoietin-1-Induced Endothelial Cell Proliferation, Migration, and Differentiation

Critical Care Division, Department of Medicine, McGill University Health Centre, McGill University, Montreal, Québec, Canada.
Arteriosclerosis Thrombosis and Vascular Biology (Impact Factor: 6). 02/2009; 29(2):209-16. DOI: 10.1161/ATVBAHA.108.181073
Source: PubMed


Angiopoietin-1 (Ang-1) is an important regulator of angiogenesis in endothelial cells. It promotes migration, proliferation, and differentiation of cells, although the regulating factors involved in these processes remain unclear. In this study, we evaluated the contribution of the transcription factor early growth response-1 (Egr-1) to Ang-1-induced angiogenesis in human umbilical vein endothelial cells (HUVECs).
Expression of Egr-1 was evaluated with real-time PCR and immunoblotting, whereas Egr-1 DNA binding activity was monitored with electrophoretic mobility shift assays. Cell migration was measured with wound healing and Boyden chamber assays, whereas cell proliferation and differentiation of cells into capillary-like tube structures were monitored with cell counting, BrdU incorporation and Matrigels. To selectively inhibit Egr-1 expression, we used both siRNA oligonucleotides and specific DNAzymes. Egr-1 mRNA expression rose approximately 9-fold within 2 hours of Ang-1 exposure and declined thereafter. Upregulation of Egr-1 expression was accompanied by an increase in nuclear mobilization and augmented DNA binding. These processes were mediated through the Erk1/2, PI-3 kinase/AKT, and mTOR pathways. Knockdown of Egr-1 expression completely abrogated Ang-1-induced endothelial migration and significantly reduced proliferation and capillary-like tube formation of HUVECs that overexpress Ang-1.
Ang-1 triggers significant and transient induction of Egr-1, and Egr-1 contributes to Ang-1-induced endothelial cell migration and proliferation.

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Available from: Nelly A Abdel-Malak, Nov 05, 2014
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    • "These findings suggest that Egr‐1 is involved in the regulation of DUSP1 expression in HUVECs exposed to Ang‐1.22 Recently, we have observed that Egr‐1 expression and activity are rapidly induced in HUVECs exposed to Ang‐1 and that Egr‐1 is essential to the proangiogenic effects of Ang‐1 in ECs.18 DUSP1 is a transcriptional target of several factors including p53, c‐Jun, ATF2, CREB, E2F, and SAP‐1.23–26 To our knowledge, no information is as yet available regarding Egr‐1 regulation of DUSP1 expression. "
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    ABSTRACT: Angiopoietin-1 (Ang-1) promotes survival and migration of endothelial cells, in part through the activation of mitogen-activated protein kinase (MAPK) pathways downstream of Tie-2 receptors. Dual-specificity phosphatases (DUSPs) dephosphorylate phosphotyrosine and phosphoserine/phosphothreonine residues on target MAPKs. The mechanisms by which DUSPs modulate MAPK activation in Ang-1/Tie-2 receptor signaling are unknown in endothelial cells. Expression of various DUSPs in human umbilical vein endothelial cells exposed to Ang-1 was measured. The functional roles of DUSPs in Ang-1-induced regulation of MAPK activation, endothelial cell survival, migration, differentiation, and permeability were measured using selective siRNA oligos. Ang-1 differentially induces DUSP1, DUSP4, and DUSP5 in human umbilical vein endothelial cells through activation of the PI-3 kinase, ERK1/2, p38, and SAPK/JNK pathways. Lack-of-function siRNA screening revealed that DUSP1 preferentially dephosphorylates p38 protein and is involved in Ang-1-induced cell migration and differentiation. DUSP4 preferentially dephosphorylates ERK1/2, p38, and SAPK/JNK proteins and, under conditions of serum deprivation, is involved in Ang-1-induced cell migration, several antiapoptotic effects, and differentiation. DUSP5 preferentially dephosphorylates ERK1/2 proteins and is involved in cell survival and inhibition of permeability. DUSP1, DUSP4, and DUSP5 differentially modulate MAPK signaling pathways downstream of Tie-2 receptors, thus highlighting the importance of these phosphatases to Ang-1-induced angiogenesis.
    Full-text · Article · Oct 2013 · Journal of the American Heart Association
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    • "This results from Angiopoietin-1 induced dephosphorylation of PECAM-1 and VE-Cadherin, respectively [20]. Endothelial cell migration, proliferation, and differentiation on the other hand are also stimulated by the protein [21]. Aim of the study was therefore to investigate the effects of Angiopoietin-1 in the setting of an eEOC-based therapy of iAKI. "
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    ABSTRACT: Acute kidney injury (AKI) severely worsens prognosis of hospitalized patients. Early Endothelial Outgrowth Cells act protective in murine acute ischemic renal failure and renoprotective actions of eEOCs have been documented to increase after cell pretreatment with 8-O-cAMP and Melatonin. Angiopoietin-1 is critically involved in maintaining vascular integrity and regeneration. Aim of the study was to analyze the consequences of eEOC treatment with Ang-1 in murine AKI. After 40 minutes of unilateral renal artery clamping with contralateral nephrectomy, male C57/Bl6N mice were injected with either untreated or pretreated (Ang-1) syngeneic murine eEOCs. Two days later serum creatinine levels and morphology were evaluated. Cultured, Ang-1 treated murine eEOCs were analyzed for production / release of proangiogenic and proinflammatory mediators, migratory activity, and cell survival, respectively. Angiopoietin-1 pretreatment of eEOCs significantly reduced serum creatinine in cell-injected mice. In vitro analysis showed increased migration of Ang-1 treated eEOCs and supernatant from Ang-1 treated eEOCs stimulated migration of cultured mature endothelial cells. In addition, Ang-1 reduced percentages of Annexin V+/PI+ eEOCs. Intrarenal numbers of eEOCs remained unaffected by Ang-1 and eEOCs did not produce more or less proangiogenic / proinflammatory mediators after being stimulated with Ang-1. Angiopoietin-1 pretreatment of eEOCs increases the cells' renoprotective competence in ischemic AKI. Thus, the armentarium of eEOC agonists in AKI is increasingly being expanded and the treatment of AKI with eEOCs becomes a promising future option.
    Full-text · Article · Oct 2013 · BMC Nephrology
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    • "The observed enrichment of this gene in cortical migrating interneurons suggests that it may positively control interneuron migration and/or differentiation. Accordingly, angiopoietin-1 (Ang-1) was found to stimulate cell migration in endothelial cells by increasing expression of Egr1 (Abdel-Malak et al., 2009). In addition, it has been reported that Reelin, produced by the mitral cells of the olfactory bulb (OB), activates the MAPK/extracellular signal-regulated kinase (ERK) pathway and induces Egr-1 transcription, which in turn promotes the shift from tangential/chain to radial/individual neuronal migration, and the detachment of neurons in the rostral migratory stream (RMS; Simó et al., 2007). "
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    ABSTRACT: Mutations in the homeobox transcription factor ARX have been found to be responsible for a wide spectrum of disorders extending from phenotypes with severe neuronal migration defects, such as lissencephaly, to mild forms of intellectual disabilities without apparent brain abnormalities, but with associated features of dystonia and epilepsy. Arx expression is mainly restricted to populations of GABA-containing neurons. Studies of the effects of ARX loss of function, either in humans or mutant mice, revealed varying defects, suggesting multiple roles of this gene in brain patterning, neuronal proliferation and migration, cell maturation and differentiation, as well as axonal outgrowth and connectivity. However, to date, little is known about how Arx functions as a transcription factor or which genes it binds and regulates. Recently, we combined chromatin immunoprecipitation and mRNA expression with microarray analysis and identified approximately 1000 gene promoters bound by Arx in transfected neuroblastoma N2a cells and mouse embryonic brain. To narrow the analysis of Arx targets to those most likely to control cortical interneuron migration and/or differentiation, we compare here our data to previously published studies searching for genes enriched or down-regulated in cortical interneurons between E13.5 and E15.5. We thus identified 14 Arx-target genes enriched (Cxcr7, Meis1, Ppap2a, Slc 12a5, Ets2, Phlda1, Egr1, Igf1, Lmo3, Sema6, Lgi1, Alk, Tgfb3, and Napb) and 5 genes specifically down-regulated (Hmgn3, Lmo1, Ebf3, Rasgef1b, and Slit2) in cortical migrating neurons. In this review, we present these genes and discuss how their possible regulation by Arx may lead to the dysfunction of GABAergic neurons, resulting in mental retardation and epilepsy.
    Full-text · Article · Dec 2011 · Frontiers in Cellular Neuroscience
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