title: A phosphoinositide 3-kinase/phospholipase Cgamma1 pathway regulates fibroblast growth factor-induced capillary tube formation.
abstract: BACKGROUND
  The fibroblast growth factors (FGFs) are key regulators of embryonic development, tissue homeostasis and tumour angiogenesis. Binding of FGFs to their receptor(s) results in activation of several intracellular signalling cascades including phosphoinositide 3-kinase (PI3K) and phospholipase C (PLC)gamma1. Here we investigated the basic FGF (FGF-2)-mediated activation of these enzymes in human umbilical vein endothelial cells (HUVECs) and defined their role in FGF-2-dependent cellular functions.
  
  
  RESULTS
  We show that FGF-2 activates PLCgamma1 in HUVECs measured by analysis of total inositol phosphates production upon metabolic labelling of cells and intracellular calcium increase. We further demonstrate that FGF-2 activates PI3K, assessed by analysing accumulation of its lipid product phosphatidylinositol-3,4,5-P(3) using TLC and confocal microscopy analysis. PI3K activity is required for FGF-2-induced PLCgamma1 activation and the PI3K/PLCgamma1 pathway is involved in FGF-2-dependent cell migration, determined using Transwell assay, and in FGF-2-induced capillary tube formation (tubulogenesis assays in vitro). Finally we show that PI3K-dependent PLCgamma1 activation regulates FGF-2-mediated phosphorylation of Akt at its residue Ser473, determined by Western blotting analysis. This occurs through protein kinase C (PKC)alpha activation since dowregulation of PKCalpha expression using specific siRNA or blockade of its activity using chemical inhibition affects the FGF-2-dependent Ser473 Akt phosphorylation. Furthermore inhibition of PKCalpha blocks FGF-2-dependent cell migration.
  
  
  CONCLUSIONS
  These data elucidate the role of PLCgamma1 in FGF-2 signalling in HUVECs demonstrating its key role in FGF-2-dependent tubulogenesis. Furthermore these data unveil a novel role for PLCgamma1 as a mediator of PI3K-dependent Akt activation and as a novel key regulator of different Akt-dependent processes.
authors.affiliation: Queen Mary University of London, Barts and The London School of Medicine and Dentistry, Blizard Institute of Cell and Molecular Science, Centre for Diabetes, Inositide Signalling Group, London, United Kingdom.
authors.forename: Tania
authors.initials: T
authors.lastname: Maffucci
authors.forename: Claudio
authors.initials: C
authors.lastname: Raimondi
authors.forename: Shadi
authors.initials: S
authors.lastname: Abu-Hayyeh
authors.forename: Veronica
authors.initials: V
authors.lastname: Dominguez
authors.forename: Gianluca
authors.initials: G
authors.lastname: Sala
authors.forename: Ian
authors.initials: I
authors.lastname: Zachary
authors.forename: Marco
authors.initials: M
authors.lastname: Falasca
mesh_terms: D002196:Capillaries / Q000187:drug effects* / Q000201:enzymology*; D002465:Cell Movement / Q000187:drug effects; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology*; D006801:Humans; D018919:Neovascularization, Physiologic / Q000187:drug effects*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D051966:Phospholipase C gamma / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D051571:Protein Kinase C-alpha / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D014471:Umbilical Veins / Q000166:cytology
chemical_list: D016222:Fibroblast Growth Factor 2; D019869:Phosphatidylinositol 3-Kinases; D051571:Protein Kinase C-alpha; D051966:Phospholipase C gamma