Early growth response-1 regulates angiopoietin-1-induced endothelial cell proliferation, migration, and differentiation.
OBJECTIVE
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).


RESULTS
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.


CONCLUSIONS
Ang-1 triggers significant and transient induction of Egr-1, and Egr-1 contributes to Ang-1-induced endothelial cell migration and proliferation.
Abdel-Malak
Nelly A
NA
Critical Care Division, Department of Medicine, McGill University Health Centre, McGill University, Montreal, Québec, Canada.
Mofarrahi
Mahroo
M
Mayaki
Dominique
D
Khachigian
Levon M
LM
Hussain
Sabah N A
SN
D042683:Angiopoietin-1 / Q000378:metabolism*; D002454:Cell Differentiation; D002465:Cell Movement; D049109:Cell Proliferation; D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000378:metabolism; D051766:Early Growth Response Protein 1 / Q000235:genetics / Q000378:metabolism*; D042783:Endothelial Cells / Q000201:enzymology / Q000378:metabolism*; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D018919:Neovascularization, Physiologic; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction; D058570:TOR Serine-Threonine Kinases; D013997:Time Factors; D014162:Transfection; D015854:Up-Regulation; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
D042683:Angiopoietin-1; D004247:DNA; D004277:DNA, Single-Stranded; D051766:Early Growth Response Protein 1; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D019869:Phosphatidylinositol 3-Kinases; D011494:Protein Kinases; D051057:Proto-Oncogene Proteins c-akt; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D058570:TOR Serine-Threonine Kinases; D042461:Vascular Endothelial Growth Factor A; C515576:COMP-Ang1 fusion protein; C400152:DNA enzyme ED5; C497325:EGR1 protein, human; C546842:MTOR protein, human