Pre-bled-young-rats in genotoxicity testing: a model for peripheral blood micronucleus assay.
Previously we have reported that prior bleeding increases the sensitivity of micronucleus (MN) assay in rats (Vikram et al., 2007 a). Rat peripheral blood micronucleus (PBMN) assay is generally not considered as reliable method for the assessment of clastogenic potential of test chemicals due to selective elimination of micronucleated cells from the circulation. The present study is aimed to evaluate the sensitivity of pre-bled-young-rat model in detecting genotoxins having different mechanism of action. In the present study, young male Sprague-Dawley (SD) rats (21-24 days old, weighing 60+/-5 g) and swiss mice (24-28 days old, weighing 15+/-2g) were used. Streptozotocin (STZ, 50mg/kg), Methotrexate (MTX, 10mg/kg), N-nitrosodiethylamine (DEN, 200mg/kg), Quercetin (QC, 50mg/kg) and Zidovudine (AZT, 400mg/kg) were used in the present experiment. Effect of prior bleeding time (0, 2, 6, 12 and 24h) on the kinetics of MN formation with STZ and AZT was studied and 36 h post chemical exposure was found to be the most suitable time point for sample collection if prior bleeding time was 0, 2 and 6h. Further, the impact of prior bleeding (2h) on the kinetics of MN formation in the bone marrow was evaluated with STZ and maximum MN frequency was observed after 24h. The area under curve (AUC) analysis proves that prior bleeding leads to significant increase in the incidence of micronucleated reticulocytes (RETs) in the peripheral blood as compared to respective non-bled controls. Out of five tested chemicals AZT and STZ induced significant increase in the MN frequency in non-bled animals while at the same dose MTX, AZT, QC and STZ induced significant increase in MN frequency in the pre-bled-young-rats employing PBMN assay as the end point. Positive results with MTX, AZT, QC, STZ and negative results with DEN demonstrate both the sensitivity and specificity of pre-bled-young-rat model in the screening of chemicals for genotoxicity using PBMN assay as the end point.
Vikram
A
A
Department of Pharmacology and Toxicology, National Institute of Pharmaceutical Education and Research (NIPER), Sector-67, SAS Nagar, Mohali, Chandigarh, Punjab 160 062, India.
Tripathi
D N
DN
Pawar
A A
AA
Ramarao
P
P
Jena
G B
GB
D000818:Animals; D019540:Area Under Curve; D004052:Diethylnitrosamine / Q000633:toxicity; D004195:Disease Models, Animal; D006470:Hemorrhage; D008297:Male; D008727:Methotrexate / Q000633:toxicity; D051379:Mice; D015162:Micronucleus Tests / Q000379:methods; D009152:Mutagenicity Tests / Q000379:methods; D009153:Mutagens / Q000633:toxicity*; D011794:Quercetin / Q000633:toxicity; D051381:Rats; D017207:Rats, Sprague-Dawley; D012156:Reticulocytes / Q000378:metabolism; D013311:Streptozocin / Q000633:toxicity; D013997:Time Factors; D015215:Zidovudine / Q000633:toxicity
D009153:Mutagens; D004052:Diethylnitrosamine; D015215:Zidovudine; D013311:Streptozocin; D011794:Quercetin; D008727:Methotrexate