title: Caffeine Suppresses Apoptosis of Bladder Cancer RT4 Cells in Response to Ionizing Radiation by Inhibiting Ataxia Telangiectasia Mutated-Chk2-p53 Axis.
abstract: BACKGROUND
  Caffeine suppresses ataxia telangiectasia and Rad3 related and ataxia telangiectasia mutated (ATM) activities; ATM is the major kinase for DNA damage detection. This study aimed to investigate the effects of caffeine on DNA damage responses in cells from the bladder cancer cell line RT4 those were exposed to ionizing radiation (IR).
  
  
  METHODS
  Immunofluorescent staining was performed to investigate changes in the proteins involved in DNA damage responses with or without caffeine. A mouse xenograft model was used to study the effects of caffeine on the DNA damage responses. Western blotting was used to investigate the effects of caffeine pretreatment on the ATM-Chk2-p53-Puma axis, while real-time polymerase chain reaction (RT-PCR) assessed changes in messenger RNA levels of p53 and downstream targets responding to IR. Finally, terminal deoxynucleotidyl transferase-dUTP nick end labeling assay. Western blotting and colony formation assay were used to measure the effects of caffeine on radiation-related apoptosis. All of the data were analyzed with a two-tailed Student's t-test.
  
  
  RESULTS
  Immunofluorescent staining showed that caffeine pretreatment profoundly suppressed the formation of γH2AXand p53-binding protein 1 foci in RT4 cells in response to irradiation. Cellular and animal experiments suggested that this suppression was mediated by suppression of the ATM-Chk2-p53-Puma DNA damage-signaling axis. RT-PCR indicated caffeine also attenuated transactivation of p53 and p53-inducible genes. The colony formation assay revealed that caffeine displayed radioprotective effects on RT4 cells in response to low-dose radiation compared to the radiosensitization effects on T24 cells.
  
  
  CONCLUSIONS
  Caffeine may inhibit IR-related apoptosis of bladder cancer RT4 cells by suppressing activation of the ATM-Chk2-p53-Puma axis.
authors.affiliation: Department of Urology, Second Affiliated Hospital; Ministry of Education Key Laboratory of Cancer Prevention and Intervention, Zhejiang University School of Medicine, Hangzhou, Zhejiang 310009, China.
authors.forename: Zhe-Wei
authors.initials: ZW
authors.lastname: Zhang
authors.forename: Jing
authors.initials: J
authors.lastname: Xiao
authors.forename: Wei
authors.initials: W
authors.lastname: Luo
authors.forename: Bo-Han
authors.initials: BH
authors.lastname: Wang
authors.forename: Ji-Min
authors.initials: JM
authors.lastname: Chen
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000528:radiation effects; D002110:Caffeine / Q000494:pharmacology*; D018797:Cell Cycle Proteins / Q000378:metabolism*; D045744:Cell Line, Tumor; D064447:Checkpoint Kinase 2 / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D051379:Mice; D008819:Mice, Nude; D011839:Radiation, Ionizing; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D001749:Urinary Bladder Neoplasms / Q000532:radiotherapy*
chemical_list: D018797:Cell Cycle Proteins; D016159:Tumor Suppressor Protein p53; D002110:Caffeine; D064447:Checkpoint Kinase 2; C578633:Chek2 protein, mouse