title: Protease-mediated human smooth muscle cell proliferation by urokinase requires epidermal growth factor receptor transactivation by triple membrane signaling.
abstract: BACKGROUND
  Urokinase (uPA) modulates cellular and extracellular matrix responses within the microenvironment of the vessel wall and has been shown to activate the epidermal growth factor receptor (EGFR). This study examines the role of the protease domain of uPA during EGFR activation in human vascular smooth muscle cells (VSMC).
  
  
  METHODS
  Human coronary VSMC were cultured in vitro. Assays of cell proliferation and EGFR phosphorylation were examined in response to the carboxyterminal fragment of uPA (CTF) in the presence and absence of the plasmin, metalloprotease and a disintegrin and metalloproteinase (ADAM) inhibitors, heparin-bound epidermal growth factor (HB-EGF), and EGFR inhibitors, and small interfering RNA to EGFR and ADAMs.
  
  
  RESULTS
  CTF produced a dose-dependent increase in DNA synthesis and cell proliferation in human VSMC, which was blocked in a dose-dependent manner by both plasmin inhibitors and the EGFR inhibitor, AG1478. CTF induced time-dependent EGFR phosphorylation, which was blocked by inhibitors of plasmin and metalloproteinases activity. The presence of urokinase plasminogen activator receptor was not required. Inhibition of ADAM-10 and -12, and of HB-EGF blocked EGFR activation in response to CTF. CTF-mediated activation of EGFR was mediated through Gβγ, src, and NAD(P)H oxidase.
  
  
  CONCLUSIONS
  In human coronary VSMC, uPA induces uPAR-independent, domain-dependent smooth muscle cell proliferation through transactivation of EGFR by a plasmin-mediated, ADAM-induced, and HB-EGF-dependent process, which is mediated by the intracellular pathways involving Gαi, Gβγ, src, and NAD(P)H oxidase.
authors.affiliation: Vascular Biology and Therapeutics Program, Houston Methodist Research Institute, Houston Methodist Hospital, Houston, Texas.
authors.forename: Enrico A
authors.initials: EA
authors.lastname: Duru
authors.affiliation: Vascular Biology and Therapeutics Program, Houston Methodist Research Institute, Houston Methodist Hospital, Houston, Texas.
authors.forename: Yuyang
authors.initials: Y
authors.lastname: Fu
authors.affiliation: Vascular Biology and Therapeutics Program, Houston Methodist Research Institute, Houston Methodist Hospital, Houston, Texas; Department of Cardiovascular Surgery, Houston Methodist DeBakey Heart & Vascular Center, Houston Methodist Hospital, Houston, Texas. Electronic address: mdavies@tmhs.org.
authors.forename: Mark G
authors.initials: MG
authors.lastname: Davies
mesh_terms: D051722:ADAM Proteins / Q000378:metabolism; D000072197:ADAM10 Protein; D000072199:ADAM12 Protein; D053829:Amyloid Precursor Protein Secretases / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000166:cytology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology*; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D010766:Phosphorylation / Q000187:drug effects / Q000502:physiology; D017434:Protein Structure, Tertiary; D011799:Quinazolines / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000235:genetics; D055293:Receptors, Urokinase Plasminogen Activator / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D020032:Tyrphostins / Q000494:pharmacology; D014568:Urokinase-Type Plasminogen Activator / Q000737:chemistry / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D008565:Membrane Proteins; D011799:Quinazolines; D034741:RNA, Small Interfering; D055293:Receptors, Urokinase Plasminogen Activator; D020032:Tyrphostins; C101044:RTKI cpd; D066246:ErbB Receptors; D053829:Amyloid Precursor Protein Secretases; D014568:Urokinase-Type Plasminogen Activator; D051722:ADAM Proteins; D000072199:ADAM12 Protein; C000606480:ADAM12 protein, human; D000072197:ADAM10 Protein; C491236:ADAM10 protein, human
keywords: Cell signaling; Epidermal growth factor receptor; Human coronary smooth muscle cell; Proliferation; Protease domain; uPA
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