title: Effects of mild temperature hyperthermia and p53 status on the size of hypoxic fractions in solid tumors, with reference to the effect in intratumor quiescent cell populations.
abstract: OBJECTIVE
  To determine the effects of mild temperature hyperthermia (MTH) and p53 status of tumor cells on the size of hypoxic fractions (HFs) in solid tumors, with reference to the effect on intratumor quiescent (Q) cell populations.
  
  
  METHODS
  Human head-and-neck squamous cell carcinoma cells transfected with mutant TP53 (SAS/mp53) or with neo vector as a control (SAS/neo) were inoculated subcutaneously into left hind legs of Balb/cA nude mice. Mice bearing the tumors received 5-bromo-2'-deoxyuridine (BrdU) continuously to label all proliferating (P) cells in the tumors. The mice then received nicotinamide injection or carbogen gas (95% O(2), 5% CO(2)) inhalation combined with or without MTH. Nicotinamide prevents intermittent blood flow that could induce perfusion-limited acute hypoxia. Chronically hypoxic cells in regions beyond the limitation of oxygen diffusion in tumors are oxygenated by increasing the oxygen transport capacity of circulating blood with carbogen gas inhalation. After each treatment, the mice received a series of test doses of gamma-rays while alive or after tumor clamping to obtain HFs in the tumors. Immediately after irradiation, the tumors were excised, minced, and trypsinized. The tumor cell suspensions thus obtained were incubated with a cytokinesis blocker (cytochalasin-B) to inhibit cytoplasmic division while allowing nuclear division. Tumor cells not labeled with BrdU were detected with immunofluorescence staining of BrdU for P cells, and the micronucleus frequency in cells without BrdU labeling [ = Q cells] was determined. The micronucleus frequency in total (P + Q) tumor cells was determined from the tumors that were not pretreated with BrdU.
  
  
  RESULTS
  SAS/mp53 tumors showed larger values for the size of not only the HF but also the diffusion-limited chronically HF than SAS/neo tumors. Q cell populations included a larger HF, particularly the chronically HF, than total cell populations in both tumors, especially in SAS/neo tumors. MTH could efficiently oxygenate the chronically HF, irrespective of p53 status.
  
  
  CONCLUSIONS
  MTH is a useful combined treatment with a radioenhancement effect on intratumor Q cells, irrespective of the p53 status of tumor cells. The p53 status has the potential to affect microenvironmental conditions within solid tumors.
authors.affiliation: Radiation Oncology Research Laboratory, Research Reactor Institute, Kyoto, University, Osaka, Japan. smasuna@rri.kyoto-u.ac.jp
authors.forename: Shin-Ichiro
authors.initials: S
authors.lastname: Masunaga
authors.forename: Akihisa
authors.initials: A
authors.lastname: Takahashi
authors.forename: Ken
authors.initials: K
authors.lastname: Ohnishi
authors.forename: Takeo
authors.initials: T
authors.lastname: Ohnishi
authors.forename: Kenji
authors.initials: K
authors.lastname: Nagata
authors.forename: Minoru
authors.initials: M
authors.lastname: Suzuki
authors.forename: Yuko
authors.initials: Y
authors.lastname: Kinashi
authors.forename: Koji
authors.initials: K
authors.lastname: Ono
mesh_terms: D000818:Animals; D002294:Carcinoma, Squamous Cell / Q000235:genetics / Q000473:pathology / Q000503:physiopathology* / Q000532:radiotherapy; D015687:Cell Hypoxia / Q000235:genetics; D016158:Genes, p53 / Q000235:genetics*; D006258:Head and Neck Neoplasms / Q000235:genetics / Q000473:pathology / Q000503:physiopathology* / Q000532:radiotherapy; D006801:Humans; D006979:Hyperthermia, Induced / Q000379:methods*; D051379:Mice; D008809:Mice, Inbred C3H; D008819:Mice, Nude; D015162:Micronucleus Tests; D009154:Mutation; D014162:Transfection