title: Sensitivity to anti-Fas is independent of increased cathepsin D activity and adrenodoxin reductase expression occurring in NOS-3 overexpressing HepG2 cells.
abstract: Stable overexpression of endothelial nitric oxide synthase (NOS-3) in HepG2 cells (4TO-NOS) leads to increased nitro-oxidative stress and upregulation of the cell death mediators p53 and Fas. Thus, NOS-3 overexpression has been suggested as a useful antiproliferative mechanism in hepatocarcinoma cells. We aimed to identify the underlying mechanism of cell death induced by NOS-3 overexpression at basal conditions and with anti-Fas treatment. The intracellular localization of NOS-3, the nitro-oxidative stress and the mitochondrial activity were analysed. In addition, the protein expression profile in 4TO-NOS was screened for differentially expressed proteins potentially involved in the induction of apoptosis. NOS-3 localization in the mitochondrial outer membrane was not associated with changes in the respiratory cellular capacity, but was related to the mitochondrial biogenesis increase and with a higher protein expression of mitochondrial complex IV. Nitro-oxidative stress and cell death in NOS-3 overexpressing cells occurred with the expression increase of pro-apoptotic genes and a higher expression/activity of the enzymes adrenodoxin reductase mitochondrial (AR) and cathepsin D (CatD). CatD overexpression in 4TO-NOS was related to the apoptosis induction independently of its catalytic activity. In addition, CatD activity inhibition by pepstatin A was not effective in blocking apoptosis induced by anti-Fas. In summary, NOS-3 overexpression resulted in an increased sensitivity to anti-Fas induced cell death, independently of AR expression and CatD activity.
authors.affiliation: Unidad de Gestión Clínica de Aparato Digestivo, Instituto Maimónides de Investigación Biomédica de Córdoba (IMIBIC)/Hospital Universitario Reina Sofía/Universidad de Córdoba, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBEREHD), Córdoba, Spain.
authors.forename: Clara I
authors.initials: CI
authors.lastname: Linares
authors.affiliation: Unidad de Gestión Clínica de Aparato Digestivo, Instituto Maimónides de Investigación Biomédica de Córdoba (IMIBIC)/Hospital Universitario Reina Sofía/Universidad de Córdoba, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBEREHD), Córdoba, Spain. Electronic address: gusfesa@gmail.com.
authors.forename: Gustavo
authors.initials: G
authors.lastname: Ferrín
authors.affiliation: Unidad de Gestión Clínica de Aparato Digestivo, Instituto Maimónides de Investigación Biomédica de Córdoba (IMIBIC)/Hospital Universitario Reina Sofía/Universidad de Córdoba, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBEREHD), Córdoba, Spain.
authors.forename: Patricia
authors.initials: P
authors.lastname: Aguilar-Melero
authors.affiliation: Unidad de Gestión Clínica de Aparato Digestivo, Instituto Maimónides de Investigación Biomédica de Córdoba (IMIBIC)/Hospital Universitario Reina Sofía/Universidad de Córdoba, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBEREHD), Córdoba, Spain.
authors.forename: Sandra
authors.initials: S
authors.lastname: González-Rubio
authors.affiliation: Unidad de Gestión Clínica de Aparato Digestivo, Instituto Maimónides de Investigación Biomédica de Córdoba (IMIBIC)/Hospital Universitario Reina Sofía/Universidad de Córdoba, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBEREHD), Córdoba, Spain.
authors.forename: Manuel
authors.initials: M
authors.lastname: Rodríguez-Perálvarez
authors.affiliation: Departamento de Biología Molecular, Centro de Biología Molecular Servero Ochoa, Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER), Centro de Investigación Hospital 12 de Octubre, ISCIII, Universidad Autónoma, Madrid, Spain.
authors.forename: María
authors.initials: M
authors.lastname: Sánchez-Aragó
authors.affiliation: Unidad de Gestión Clínica de Aparato Digestivo, Instituto Maimónides de Investigación Biomédica de Córdoba (IMIBIC)/Hospital Universitario Reina Sofía/Universidad de Córdoba, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBEREHD), Córdoba, Spain.
authors.forename: Eduardo
authors.initials: E
authors.lastname: Chicano-Gálvez
authors.affiliation: Departamento de Biología Molecular, Centro de Biología Molecular Servero Ochoa, Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER), Centro de Investigación Hospital 12 de Octubre, ISCIII, Universidad Autónoma, Madrid, Spain.
authors.forename: José M
authors.initials: JM
authors.lastname: Cuezva
authors.affiliation: Unidad de Gestión Clínica de Aparato Digestivo, Instituto Maimónides de Investigación Biomédica de Córdoba (IMIBIC)/Hospital Universitario Reina Sofía/Universidad de Córdoba, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBEREHD), Córdoba, Spain.
authors.forename: José L
authors.initials: JL
authors.lastname: Montero-Álvarez
authors.affiliation: Unidad de Gestión Clínica de Aparato Digestivo, Instituto Maimónides de Investigación Biomédica de Córdoba (IMIBIC)/Hospital Universitario Reina Sofía/Universidad de Córdoba, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBEREHD), Córdoba, Spain.
authors.forename: Jordi
authors.initials: J
authors.lastname: Muntané
authors.affiliation: Unidad de Gestión Clínica de Aparato Digestivo, Instituto Maimónides de Investigación Biomédica de Córdoba (IMIBIC)/Hospital Universitario Reina Sofía/Universidad de Córdoba, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBEREHD), Córdoba, Spain.
authors.forename: Manuel
authors.initials: M
authors.lastname: de la Mata
mesh_terms: D002402:Cathepsin D / Q000378:metabolism*; D016923:Cell Death; D019069:Cell Respiration; D004272:DNA, Mitochondrial / Q000235:genetics; D005287:Ferredoxin-NADP Reductase / Q000378:metabolism*; D018628:Gene Dosage; D056945:Hep G2 Cells; D006801:Humans; D051336:Mitochondrial Membranes / Q000378:metabolism; D063269:Mitochondrial Turnover; D008954:Models, Biological; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D010085:Oxidative Phosphorylation; D018384:Oxidative Stress; D021381:Protein Transport; D020543:Proteome / Q000378:metabolism; D040901:Proteomics; D019014:fas Receptor / Q000378:metabolism*
chemical_list: D004272:DNA, Mitochondrial; C491901:FAS protein, human; D020543:Proteome; D019014:fas Receptor; C496314:NOS3 protein, human; D052250:Nitric Oxide Synthase Type III; D005287:Ferredoxin-NADP Reductase; D002402:Cathepsin D
keywords: Adrenodoxin reductase; Cathepsin D; Endothelial nitric oxide synthase; Fas-mediated apoptosis; Hepatocarcinoma; Nitro-oxidative stress