title: Increased oxidative DNA damage in mammary tumor cells by continuous epidermal growth factor stimulation.
abstract: BACKGROUND
  Growth factors can enhance the malignant potential of tumor cells. To examine the relationship between growth factors and tumor progression, we previously established a weakly malignant cell line, ER-1. We found that a 24-hour exposure of ER-1 cells to epidermal growth factor (EGF) induced malignant properties (tumor progression) that were reversible but that, after a 1-month exposure, these changes were irreversible. In this study, we investigated the irreversible changes induced in ER-1 cells by a 1-month exposure to EGF and the possible involvement of oxidative stress.
  
  
  METHODS
  ER-1 cells were treated with EGF (100 ng/mL) for 1 month in the presence or absence of an antioxidant, N-acetylcysteine or selenium, and compared with untreated control ER-1 cells. We assessed tumor progression by measuring intracellular peroxide levels, 8-hydroxydeoxyguanosine (a marker for oxidative DNA damage) levels, in vitro invasiveness, and in vivo tumorigenicity and metastatic ability. All statistical tests are two-sided.
  
  
  RESULTS
  After ER-1 cells were treated for 1 month with EGF, levels of intracellular peroxide and 8-hydroxyguanosine in the DNA of treated cells were higher than those in the DNA of control cells, and treated ER-1 cells were more tumorigenic and metastatic in vivo and more invasive in vitro than untreated control cells (all P<.001). Levels of 8-hydroxyguanosine in DNA increased as the length of the EGF treatment increased (P<.001). However, when N-acetylcysteine or selenium was added with EGF for 1 month, levels of intracellular peroxide and 8-hydroxyguanosine in DNA were comparable to those in control cells (r =.795). Both tumorigenicity (P =.008) and metastatic ability (P<.001) decreased after addition of N-acetylcysteine or selenium.
  
  
  CONCLUSIONS
  The irreversible changes caused by continuous EGF stimulation of ER-1 cells result from increased oxidative damage in the DNA, which generates tumor cells with more malignant characteristics.
authors.affiliation: Division of Cancer-Related Genes, Institute for Genetic Medicine, Hokkaido University, Sapporo, Japan. jhamada@med.hokudai.ac.jp
authors.forename: J
authors.initials: J
authors.lastname: Hamada
authors.forename: D
authors.initials: D
authors.lastname: Nakata
authors.forename: D
authors.initials: D
authors.lastname: Nakae
authors.forename: Y
authors.initials: Y
authors.lastname: Kobayashi
authors.forename: H
authors.initials: H
authors.lastname: Akai
authors.forename: Y
authors.initials: Y
authors.lastname: Konishi
authors.forename: F
authors.initials: F
authors.lastname: Okada
authors.forename: T
authors.initials: T
authors.lastname: Shibata
authors.forename: M
authors.initials: M
authors.lastname: Hosokawa
authors.forename: T
authors.initials: T
authors.lastname: Moriuchi
mesh_terms: D000080242:8-Hydroxy-2'-Deoxyguanosine; D000111:Acetylcysteine / Q000494:pharmacology; D000230:Adenocarcinoma / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D004249:DNA Damage / Q000187:drug effects; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000378:metabolism; D018450:Disease Progression; D004815:Epidermal Growth Factor / Q000009:adverse effects*; D005260:Female; D016166:Free Radical Scavengers / Q000494:pharmacology*; D005979:Glutathione Peroxidase / Q000378:metabolism; D008325:Mammary Neoplasms, Experimental / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism*; D018613:Microscopy, Confocal; D018384:Oxidative Stress / Q000187:drug effects; D051381:Rats; D011918:Rats, Inbred SHR; D012643:Selenium / Q000494:pharmacology; D014407:Tumor Cells, Cultured
chemical_list: D000975:Antioxidants; D016166:Free Radical Scavengers; D004815:Epidermal Growth Factor; D000080242:8-Hydroxy-2'-Deoxyguanosine; D005979:Glutathione Peroxidase; D003849:Deoxyguanosine; D012643:Selenium; D000111:Acetylcysteine