Differential activation of multiple signalling pathways dictates eNOS upregulation by FGF2 but not VEGF in placental artery endothelial cells.
Fibroblast growth factor (FGF2), but not vascular endothelial growth factor (VEGF), upregulates endothelial nitric oxide synthase (eNOS) protein expression, at least partially, via activation of extracellular signal-regulated kinase 2/1 (ERK2/1) in ovine fetoplacental artery endothelial (oFPAE) cells. Herein we further investigated the temporal effects of FGF2 and VEGF on other signalling pathways including members (Jun N-terminal kinase JNK1/2 and p38MAPK) of mitogen-activated protein kinases (MAPK), phosphatidylinositol-3 kinase/v-akt murine thymoma viral oncogene homologue 1 (PI3K/AKT1), and the tyrosine kinase c-SRC, and examined if either one or more of these pathways play a role in the differential regulation of eNOS by FGF2 and VEGF. We first confirmed that in oFPAE cells, FGF2, but not VEGF, increased eNOS protein. FGF2 stimulated eNOS protein in a time- and concentration-dependent manner, which also depended on cell density. FGF2 provoked sustained (5min to 12h) whereas VEGF only stimulated transient (5min) ERK2/1 phosphorylation. FGF2 was 1.7-fold more potent in stimulating ERK2/1 phosphorylation than VEGF. FGF2 and VEGF only transiently activated JNK1/2 and AKT1 within 5min; however, FGF2 was a stronger stimulus than VEGF. FGF2 and VEGF did not significantly activate p38MAPK at 5min; however, VEGF stimulated p38MAPK phosphorylation at 60min. VEGF but not FGF2 significantly stimulated c-SRC phosphorylation. Inhibitors of MEK-ERK2/1 (PD98059), JNK1/2 (SP600125) and PI3K (wortmannin), but not p38MAPK (SB203580) and SRC (PP2), decreased the FGF2-increased eNOS protein expression. Thus, the FGF2-induced eNOS protein expression requires activation of multiple signalling pathways including ERK2/1, JNK1/2 and PI3K/AKT1. Differences in intensity and temporal patterns of activation of these pathways by FGF2 and VEGF may account for their differential effects on eNOS expression in OFPAE cells.
Division of Maternal-Fetal Medicine (MC0802), Department of Reproductive Medicine, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0802, USA.
Eugenia
E
Mata-Greenwood
Wu-Xiang
WX
Liao
Jing
J
Zheng
Dong-Bao
DB
Chen
D000818:Animals; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D005260:Female; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D048055:Mitogen-Activated Protein Kinase 8 / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010920:Placenta / Q000098:blood supply / Q000187:drug effects* / Q000378:metabolism; D021041:Placental Circulation / Q000187:drug effects / Q000502:physiology; D011247:Pregnancy; D011494:Protein Kinases / Q000378:metabolism; D012756:Sheep; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D014469:Umbilical Arteries / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects*; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology*
D042461:Vascular Endothelial Growth Factor A; D016222:Fibroblast Growth Factor 2; D052250:Nitric Oxide Synthase Type III; D011494:Protein Kinases; D019869:Phosphatidylinositol 3-Kinases; D048052:Mitogen-Activated Protein Kinase 3; D048055:Mitogen-Activated Protein Kinase 8