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Dose dependent and divergent effects of superoxide anion on cell death, proliferation, and migration of activated human hepatic stellate cells
  1. E Novo1,
  2. F Marra2,
  3. E Zamara1,
  4. L Valfrè di Bonzo1,
  5. A Caligiuri2,
  6. S Cannito1,
  7. C Antonaci1,
  8. S Colombatto1,
  9. M Pinzani2,
  10. M Parola1
  1. 1Dip Medicina e Oncologia Sperimentale, University of Torino, Torino, Italy
  2. 2Dip Medicina Interna-Centro di Ricerca, Trasferimento e Alta Formazione “DENOTHE”, University of Florence, Florence, Italy
  1. Correspondence to:
    Professor M Parola
    Università degli Studi di Torino, Dip Medicina e Oncologia Sperimentale, C so Raffaello 30, 10125 Torino, Italy; maurizio.parola{at}


Background and aim: Activated myofibroblast-like cells, originating from hepatic stellate cells (HSC/MFs) or other cellular sources, play a key profibrogenic role in chronic liver diseases (CLDs) that, as suggested by studies in animal models or rat HSC/MFs, may be modulated by reactive oxygen intermediates (ROI). In this study, human HSC/MFs, exposed to different levels of superoxide anion (O2•−) and, for comparison, hydrogen peroxide (H2O2), were analysed in terms of cytotoxicity, proliferative response, and migration.

Methods: Cultured human HSC/MFs were exposed to controlled O2•− generation by hypoxanthine/xanthine oxidase systems or to a range of H2O2 concentrations. Induction of cell death, proliferation, and migration were investigated using morphology, molecular biology, and biochemical techniques.

Results: Human HSC/MFs were shown to be extremely resistant to induction of cell death by O2•− and only high rates of O2•− generation induced either necrotic or apoptotic cell death. Non-cytotoxic low levels of O2•−, able to upregulate procollagen type I expression (but not tissue inhibitor of metalloproteinase 1 and 2), stimulated migration of human HSC/MFs in a Ras/extracellular regulated kinase (ERK) dependent, antioxidant sensitive way, without affecting basal or platelet derived growth factor (PDGF) stimulated cell proliferation. Non-cytotoxic levels of H2O2 did not affect Ras/ERK or proliferative response. A high rate of O2•− generation or elevated levels of H2O2 induced cytoskeletal alterations, block in motility, and inhibition of PDGF dependent DNA synthesis.

Conclusions: Low non-cytotoxic levels of extracellularly generated O2•− may stimulate selected profibrogenic responses in human HSC/MFs without affecting proliferation.

  • CLDs, chronic liver diseases
  • ECM, extracellular matrix
  • MFs, myofibroblast-like cells
  • HSC, hepatic stellate cells
  • HSC/MFs, activated hepatic stellate cells in myofibroblast-like phenotype
  • ROI, reactive oxygen intermediates
  • O2•−, superoxide anion
  • H2O2, hydrogen peroxide
  • HNE, 4-hydroxynonenal
  • TIMP, tissue inhibitor of metalloproteinases
  • PDGF, platelet derived growth factor
  • TGF-β1, transforming growth factor β1
  • ECL, enhanced chemiluminescence
  • SFI medium, serum free Iscove’s medium
  • NO, nitric oxide
  • L-NAME, Nω-nitro-L-arginine methyl ester
  • LDH, lactate dehydrogenase
  • X/XO, hypoxanthine/xanthine oxidase system generating superoxide anion
  • SOD, superoxide dismutase
  • ERK, extracellular regulated kinase
  • PI-3 K, phosphatidyl inositol 3-kinase
  • activated human hepatic stellate cells
  • liver fibrosis
  • superoxide anion
  • myofibroblast migration
  • induction of cell death

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  • Conflict of interest: None declared.