Dihydropyrimidine dehydrogenase circadian rhythm in mouse liver: comparison between enzyme activity and gene expression.
Dihydropyrimidine dehydrogenase (DPD) is the rate-limiting enzyme of 5-fluorouracil (FU) catabolism. The relevance of the measurement of DPD activity for identifying DPD-deficient patients is lessened by circadian variability in DPD activity. Our purpose was to determine whether or not DPD mRNA is sustained by a circadian rhythm. Synchronised mice (male B6D2F1) were sacrificed at 3, 7, 11, 15, 19 or 23 Hours After Light Onset (HALO; eight mice per time-point). Liver DPD activity was determined by a radio-enzymatic assay and liver DPD expression by a reverse transcriptase-polymerase chain reaction (RT-PCR) enzyme-linked immunosorbent assay (ELISA) method. Mice synchronisation was controlled by leucocyte and neutrophil counts. Individual DPD activity ranged from 555 to 1575 pmol/min/mg prot; mean DPD activity was highest at 3 HALO (mean+/-standard error of the mean (S.E.M.); 1105+/-70) and lowest at 15 HALO (889+/-71). Individual liver DPD expression varied from 761 to 3481 units (DPD/beta actin ratio); the mean was lowest at 3 HALO (1406+/-112) and highest at 15 HALO (2067+/-214). Cosinor analysis indicated that respective double amplitudes of DPD activity and expression were 21 and 30% of the 24-h mean. The acrophases for activity and expression were 6:40 and 14:10 HALO, respectively, meaning that maximum activity occurred 16 h after the maximum observed expression. These results, revealing the existence of a circadian rhythm in DPD expression, should stimulate further studies to enhance our understanding of the molecular mechanisms involved in the circadian regulation of the DPD enzyme.
Porsin
B
B
Oncopharmacology Unit, Centre Antoine Lacassagne, Nice, France.
Formento
J-L
JL
Filipski
E
E
Etienne
M-C
MC
Francoual
M
M
Renée
N
N
Magné
N
N
Lévi
F
F
Milano
G
G
D000818:Animals; D002940:Circadian Rhythm / Q000502:physiology*; D042943:Dihydrouracil Dehydrogenase (NADP); D004797:Enzyme-Linked Immunosorbent Assay; D008099:Liver / Q000201:enzymology*; D008297:Male; D051379:Mice; D010088:Oxidoreductases / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
D012333:RNA, Messenger; D010088:Oxidoreductases; D042943:Dihydrouracil Dehydrogenase (NADP)