Simultaneous use of erythropoietin and prior bleeding enhances the sensitivity of the peripheral blood micronucleus assay

Abstract

Adult rats are generally not considered as a suitable model for the peripheral blood micronucleus (PBMN) assay in regulatory consideration, owing to the splenic removal of the micronucleated cells from circulation. Although prior bleeding (PrB) increases the sensitivity of the PBMN assay in young rats, the volume of bleeding and the associated stress caused are major concerns for its possible use in genotoxicity studies. The present study was aimed to overcome these limitations in using pre-bled young rats in genotoxicity studies. The bleeding volume was reduced by the simultaneous use of erythropoietin (EPO) to increase the sensitivity of PBMN assay. Young Sprague-Dawley (SD, 26 days) rats were used in the study. The kinetics of RETs-to-ERTs ratio was determined in response to EPO (10-3000 IU/kg) or PrB (0.1-1.0 ml) at different time points (0, 6, 12, 24, 36, 48, 72 and 96 h). Injection of EPO (30 IU/kg) and PrB (0.5 ml) led to a significant increase in the MN frequency in the PBMN assay in response to cyclophosphamide and zidovudine. The effect of EPO treatment and/or PrB on cell viability and proliferation in the bone marrow (BM) was examined. The results of the present study clearly demonstrate that the simultaneous use of both EPO and PrB enhances the sensitivity of the PBMN assay in young rats due to increased cellular proliferation in the BM. This may provide a useful experimental model for the evaluation of marginally active genotoxicants.