title: Hypoxic proliferation requires EGFR-mediated ERK activation in human pulmonary microvascular endothelial cells.
abstract: We have previously shown that hypoxic proliferation of human pulmonary microvascular endothelial cells (hPMVECs) depends on epidermal growth factor receptor (EGFR) activation. To determine downstream signaling leading to proliferation, we tested the hypothesis that hypoxia-induced proliferation in hPMVECs would require EGFR-mediated activation of extracellular signal-regulated kinase (ERK) leading to arginase II induction. To test this hypothesis, hPMVECs were incubated in either normoxia (21% O2, 5% CO2) or hypoxia (1% O2, 5% CO2) and Western blotting was performed for EGFR, arginase II, phosphorylated-ERK (pERK), and total ERK (ERK). Hypoxia led to greater EGFR, pERK, and arginase II protein levels than did normoxia in hPMVECs. To examine the role of EGFR in these hypoxia-induced changes, hPMVECs were transfected with siRNA against EGFR or a scrambled siRNA and placed in hypoxia. Inhibition of EGFR using siRNA attenuated hypoxia-induced pERK and arginase II expression as well as the hypoxia-induced increase in viable cell numbers. hPMVECs were then treated with vehicle, an EGFR inhibitor (AG1478), or an ERK pathway inhibitor (U0126) and placed in hypoxia. Pharmacologic inhibition of EGFR significantly attenuated the hypoxia-induced increase in pERK level. Both AG1478 and U0126 also significantly attenuated the hypoxia-induced increase in viable hPMVECs numbers. hPMVECs were transfected with an adenoviral vector containing arginase II (AdArg2) and overexpression of arginase II rescued the U0126-mediated decrease in viable cell numbers in hypoxic hPMVECs. Our findings suggest that hypoxic activation of EGFR results in phosphorylation of ERK, which is required for hypoxic induction of arginase II and cellular proliferation.
authors.affiliation: Pulmonary Hypertension Group, Center for Perinatal Research, Research Institute at Nationwide Children's Hospital, Columbus, Ohio; and.|Department of Pediatrics, The Ohio State University, Columbus, Ohio.
authors.forename: Hilary A
authors.initials: HA
authors.lastname: White
authors.affiliation: Pulmonary Hypertension Group, Center for Perinatal Research, Research Institute at Nationwide Children's Hospital, Columbus, Ohio; and.|Department of Pediatrics, The Ohio State University, Columbus, Ohio.
authors.forename: Yi
authors.initials: Y
authors.lastname: Jin
authors.affiliation: Pulmonary Hypertension Group, Center for Perinatal Research, Research Institute at Nationwide Children's Hospital, Columbus, Ohio; and.|Department of Pediatrics, The Ohio State University, Columbus, Ohio.
authors.forename: Louis G
authors.initials: LG
authors.lastname: Chicoine
authors.affiliation: Pulmonary Hypertension Group, Center for Perinatal Research, Research Institute at Nationwide Children's Hospital, Columbus, Ohio; and.|Department of Pediatrics, The Ohio State University, Columbus, Ohio.
authors.forename: Bernadette
authors.initials: B
authors.lastname: Chen
authors.affiliation: Pulmonary Hypertension Group, Center for Perinatal Research, Research Institute at Nationwide Children's Hospital, Columbus, Ohio; and.|Department of Pediatrics, The Ohio State University, Columbus, Ohio.
authors.forename: Yusen
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Pulmonary Hypertension Group, Center for Perinatal Research, Research Institute at Nationwide Children's Hospital, Columbus, Ohio; and leif.nelin@nationwidechildrens.org.|Department of Pediatrics, The Ohio State University, Columbus, Ohio leif.nelin@nationwidechildrens.org.
authors.forename: Leif D
authors.initials: LD
authors.lastname: Nelin
mesh_terms: D001119:Arginase / Q000378:metabolism; D002070:Butadienes / Q000494:pharmacology; D002452:Cell Count; D015687:Cell Hypoxia / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D017077:Culture Media, Conditioned / Q000494:pharmacology; D042783:Endothelial Cells / Q000187:drug effects / Q000201:enzymology*; D004789:Enzyme Activation / Q000187:drug effects; D066246:ErbB Receptors / Q000378:metabolism*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D020868:Gene Silencing / Q000187:drug effects; D006801:Humans; D008168:Lung / Q000098:blood supply*; D055806:Microvessels / Q000473:pathology*; D009570:Nitriles / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011799:Quinazolines / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000378:metabolism; D020032:Tyrphostins / Q000494:pharmacology; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: D002070:Butadienes; D017077:Culture Media, Conditioned; D009570:Nitriles; D011799:Quinazolines; D034741:RNA, Small Interfering; D020032:Tyrphostins; C113580:U 0126; C101044:RTKI cpd; D066246:ErbB Receptors; D048049:Extracellular Signal-Regulated MAP Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D001119:Arginase
keywords: arginase; hypoxia; pulmonary hypertension; pulmonary vascular remodeling