title: Delayed cell death associated with mitotic catastrophe in γ-irradiated stem-like glioma cells.
abstract: OBJECTIVE
  Stem-like tumor cells are regarded as highly resistant to ionizing radiation (IR). Previous studies have focused on apoptosis early after irradiation, and the apoptosis resistance observed has been attributed to reduced DNA damage or enhanced DNA repair compared to non-stem tumor cells. Here, early and late radioresponse of patient-derived stem-like glioma cells (SLGCs) and differentiated cells directly derived from them were examined for cell death mode and the influence of stem cell-specific growth factors.
  
  
  METHODS
  Primary SLGCs were propagated in serum-free medium with the stem-cell mitogens epidermal growth factor (EGF) and fibroblast growth factor-2 (FGF-2). Differentiation was induced by serum-containing medium without EGF and FGF. Radiation sensitivity was evaluated by assessing proliferation, clonogenic survival, apoptosis, and mitotic catastrophe. DNA damage-associated γH2AX as well as p53 and p21 expression were determined by Western blots.
  
  
  RESULTS
  SLGCs failed to apoptose in the first 4 days after irradiation even at high single doses up to 10 Gy, but we observed substantial cell death later than 4 days postirradiation in 3 of 6 SLGC lines treated with 5 or 10 Gy. This delayed cell death was observed in 3 of the 4 SLGC lines with nonfunctional p53, was associated with mitotic catastrophe and occurred via apoptosis. The early apoptosis resistance of the SLGCs was associated with lower γH2AX compared to differentiated cells, but we found that the stem-cell culture cytokines EGF plus FGF-2 strongly reduce γH2AX levels. Nonetheless, in two p53-deficient SLGC lines examined γIR-induced apoptosis even correlated with EGF/FGF-induced proliferation and mitotic catastrophe. In a line containing CD133-positive and -negative stem-like cells, the CD133-positive cells proliferated faster and underwent more γIR-induced mitotic catastrophe.
  
  
  CONCLUSIONS
  Our results suggest the importance of delayed apoptosis, associated mitotic catastrophe, and cellular proliferation for γIR-induced death of p53-deficient SLGCs. This may have therapeutic implications. We further show that the stem-cell culture cytokines EGF plus FGF-2 activate DNA repair and thus confound in vitro comparisons of DNA damage repair between stem-like and more differentiated tumor cells.
authors.affiliation: Department of Radiation Oncology, University Hospital Freiburg, Freiburg, Germany.
authors.forename: Elke
authors.initials: E
authors.lastname: Firat
authors.forename: Simone
authors.initials: S
authors.lastname: Gaedicke
authors.forename: Chizuko
authors.initials: C
authors.lastname: Tsurumi
authors.forename: Norbert
authors.initials: N
authors.lastname: Esser
authors.forename: Astrid
authors.initials: A
authors.lastname: Weyerbrock
authors.forename: Gabriele
authors.initials: G
authors.lastname: Niedermann
mesh_terms: D017209:Apoptosis; D001932:Brain Neoplasms / Q000378:metabolism / Q000473:pathology / Q000532:radiotherapy*; D049109:Cell Proliferation; D002470:Cell Survival; D004249:DNA Damage / Q000528:radiation effects; D004815:Epidermal Growth Factor / Q000378:metabolism; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism; D005720:Gamma Rays; D005910:Glioma / Q000473:pathology / Q000532:radiotherapy*; D006657:Histones / Q000378:metabolism; D006801:Humans; D008938:Mitosis / Q000528:radiation effects*; D014411:Neoplastic Stem Cells / Q000528:radiation effects*; D011836:Radiation Tolerance; D011839:Radiation, Ionizing
chemical_list: C468783:H2AX protein, human; D006657:Histones; D016222:Fibroblast Growth Factor 2; D004815:Epidermal Growth Factor