title: Stress hormones promote growth of B16-F10 melanoma metastases: an interleukin 6- and glutathione-dependent mechanism.
abstract: BACKGROUND
  Interleukin (IL)-6 (mainly of tumor origin) activates glutathione (GSH) release from hepatocytes and its interorgan transport to B16-F10 melanoma metastatic foci. We studied if this capacity to overproduce IL-6 is regulated by cancer cell-independent mechanisms.
  
  
  METHODS
  Murine B16-F10 melanoma cells were cultured, transfected with red fluorescent protein, injected i.v. into syngenic C57BL/6J mice to generate lung and liver metastases, and isolated from metastatic foci using high-performance cell sorting. Stress hormones and IL-6 levels were measured by ELISA, and CRH expression in the brain by in situ hybridization. DNA binding activity of NF-κB, CREB, AP-1, and NF-IL-6 was measured using specific transcription factor assay kits. IL-6 expression was measured by RT-PCR, and silencing was achieved by transfection of anti-IL-6 small interfering RNA. GSH was determined by HPLC. Cell death analysis was distinguished using fluorescence microscopy, TUNEL labeling, and flow cytometry techniques. Statistical analyses were performed using Student's t test.
  
  
  RESULTS
  Plasma levels of stress-related hormones (adrenocorticotropin hormone, corticosterone, and noradrenaline) increased, following a circadian pattern and as compared to non-tumor controls, in mice bearing B16-F10 lung or liver metastases. Corticosterone and noradrenaline, at pathophysiological levels, increased expression and secretion of IL-6 in B16-F10 cells in vitro. Corticosterone- and noradrenaline-induced transcriptional up-regulation of IL-6 gene involves changes in the DNA binding activity of nuclear factor-κB, cAMP response element-binding protein, activator protein-1, and nuclear factor for IL-6. In vivo inoculation of B16-F10 cells transfected with anti-IL-6-siRNA, treatment with a glucocorticoid receptor blocker (RU-486) or with a β-adrenoceptor blocker (propranolol), increased hepatic GSH whereas decreased plasma IL-6 levels and metastatic growth. Corticosterone, but not NORA, also induced apoptotic cell death in metastatic cells with low GSH content.
  
  
  CONCLUSIONS
  Our results describe an interorgan system where stress-related hormones, IL-6, and GSH coordinately regulate metastases growth.
authors.affiliation: Department of Physiology, University of Valencia, 15 Av, Blasco Ibañez, 46010 Valencia, Spain.
authors.forename: Soraya L
authors.initials: SL
authors.lastname: Valles
authors.forename: María
authors.initials: M
authors.lastname: Benlloch
authors.forename: María L
authors.initials: ML
authors.lastname: Rodriguez
authors.forename: Salvador
authors.initials: S
authors.lastname: Mena
authors.forename: José A
authors.initials: JA
authors.lastname: Pellicer
authors.forename: Miguel
authors.initials: M
authors.lastname: Asensi
authors.forename: Elena
authors.initials: E
authors.lastname: Obrador
authors.forename: José M
authors.initials: JM
authors.lastname: Estrela
mesh_terms: D000324:Adrenocorticotropic Hormone / Q000097:blood / Q000502:physiology*; D000818:Animals; D001483:Base Sequence; D045744:Cell Line, Tumor; D003345:Corticosterone / Q000097:blood / Q000502:physiology*; D015342:DNA Probes; D018274:Electroporation; D004797:Enzyme-Linked Immunosorbent Assay; D005978:Glutathione / Q000502:physiology*; D017403:In Situ Hybridization; D015850:Interleukin-6 / Q000235:genetics / Q000502:physiology*; D008546:Melanoma, Experimental / Q000473:pathology*; D051379:Mice; D009362:Neoplasm Metastasis; D009638:Norepinephrine / Q000097:blood / Q000502:physiology*; D060888:Real-Time Polymerase Chain Reaction; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000502:physiology
chemical_list: D015342:DNA Probes; D015850:Interleukin-6; D014157:Transcription Factors; D000324:Adrenocorticotropic Hormone; D005978:Glutathione; D003345:Corticosterone; D009638:Norepinephrine