Aj. Edwards et al., Lack of effect of coumarin on unscheduled DNA synthesis in the in vivo rathepatocyte DNA repair assay, FOOD CHEM T, 38(5), 2000, pp. 403-409
The ability of coumarin to induce UDS in male Spague-Dawley CD rat hepatocy
tes bt vivo was assessed using the unscheduled DNA synthesis (UDS) assay. F
rom a preliminary toxicity study the oral maximum tolerated dose (MTD) of c
oumarin was determined to be 320 mg/kg body weight. For the UDS studies, ra
ts were treated with 0 (corn oil control), 32 tone-tenth the MTD), 107 tone
-third the MTD) and 320 (MTD) mg/kg coumarin via oral gavage. Rats were als
o treated with 20 mg/kg body weight dimethylnitrosamine (DMN) or 50 mg/kg b
ody weight 2-acetylaminofluorene (2-AAF) as positive controls for the 2-4 h
r and 12-16 hr expression of UDS, respectively. Hepatocytes were isolated b
y Liver perfusion either 2-4 hr or 12-16 hr after treatment and cultured in
medium containing [methyl-H-3]-thymidine for 4 hr and assessed for UDS by
grain counting of autoradiographs. Coumarin treatment at doses of 32-320 mg
/kg body weight had no statistically significant or dose-related effect on
UDS in rat hepatocytes either 2-4 hr or 12-16 hr after dosing. In contrast,
both DMN 2-4 hr after dosing and 2-AAF 12-16 hr after dosing produced sign
ificant increases in UDS assessed as the net nuclear grain count. Both geno
toxins also increased the percentage of hepatocyte nuclei with greater than
5 net grains. Treatment with coumarin, DMN and 2-AAF had no statistically
significant effect on the proportion of rat hepatocytes undergoing replicat
ive DNA synthesis. In summary, this study demonstrates that coumarin does n
ot induce UDS in hepatocytes of male Sprague-Dawley CD rats after oral admi
nistration at doses up to the MTD of 320 mg/kg. The responsiveness of the a
nimals used in this study to genotoxic agents was demonstrated by the clear
induction of DNA repair after treatment with DMN and 2-AAF. (C) 2000 Elsev
ier Science. Ltd. All rights reserved.