title: The ability of hypoxia to modify the gene expression of thymidylate synthase in tumour cells in vivo.
abstract: OBJECTIVE
  Hypoxic cells in tumours are resistant to 5-fluorouracil (5-FU). This in vivo study investigated the ability of hypoxia to regulate the gene expression of thymidylate synthase (TS), the target enzyme of 5-FU.
  
  
  METHODS
  C3H mammary carcinomas, grown in the feet of female CDF1 mice, were used for all experiments. Mice were placed in a 10% oxygen environment for various time periods and the tumour oxygen status was determined with an Eppendorf oxygen electrode. The animals were then injected with BrdU (100 mg/kg, i.p.). Tumours were excised and immediately frozen (-80 degrees C) until isolation of total RNA. The mRNA was reversibly transcribed to complementary DNA and the resulting cDNA amplified in a multiplex PCR reaction, with beta-actin as the internal reference gene.
  
  
  RESULTS
  One hour of low oxygen breathing made tumours significantly more hypoxic. This increase was maintained for a maximum incubation period of 48 h. In the same tumours, no change in TS gene expression was seen with up to 3 h of low oxygen breathing. At longer times it decreased, reaching significance at 12-24 h and remaining at this lower level for up to 48 h. BrdU labelling was significantly reduced after breathing low O2 for 24 h (p = 0.001).
  
  
  CONCLUSIONS
  Hypoxia-induced down-regulation of TS gene expression was observed. This would be expected to make hypoxic tumour cells more sensitive to 5-FU. Other mechanisms must be responsible for the previously reported resistance to this drug.
authors.affiliation: Department of Oncology, Aarhus University Hospital, Denmark. evae@dadlnet.dk
authors.forename: E
authors.initials: E
authors.lastname: Ehrnrooth
authors.forename: H
authors.initials: H
authors.lastname: von der Maase
authors.forename: B S
authors.initials: BS
authors.lastname: Sørensen
authors.forename: J H
authors.initials: JH
authors.lastname: Poulsen
authors.forename: M R
authors.initials: MR
authors.lastname: Horsman
mesh_terms: D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D004351:Drug Resistance; D005260:Female; D005472:Fluorouracil / Q000494:pharmacology; D015870:Gene Expression; D000860:Hypoxia / Q000201:enzymology* / Q000235:genetics*; D008325:Mammary Neoplasms, Experimental / Q000201:enzymology* / Q000235:genetics* / Q000628:therapy; D051379:Mice; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012334:RNA, Neoplasm / Q000235:genetics / Q000378:metabolism; D013940:Thymidylate Synthase / Q000235:genetics*
chemical_list: D000964:Antimetabolites, Antineoplastic; D017931:DNA Primers; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D013940:Thymidylate Synthase; D005472:Fluorouracil