INDUCTION OF MICRONUCLEATED ERYTHROCYTES BY RECOMBINANT-HUMAN-ERYTHROPOIETIN

Induction of micronucleated erythrocytes by a recombinant human erythropoietin (rhEPO) was examined using both in vitro and in vivo test systems. A small, significant and dose-related increase in the frequency of micronucleated polychromatic erythrocytes (MNPCE) in bone marrow of n-mice administered i.p. with 12 500-50 000 IU/kg rhEPO was induced at 48 h sampling time. A clear positive dose-response relationship and significant increase in the frequency of micronucleated reticulocytes (MNRET) in peripheral blood of mice administered i.p. with 400-50 000 IU/kg rhEPO was noted at 48, 72 and 96 h sampling times. Conversely, in bacterial reverse mutation tests, no noticeable increase of auxotrophic revertants was observed in Salmonella typhimurium, TA100, TA98, TA1535, TA1537, or Escherichia coli, WP2 uvrA-, by treatment with 188-6000 IU/plate of rhEPO, with or without S9 mix. Furthermore, rhEPO at 750-6000 IU/ml did not induce chromosomal aberrations in vitro in CHL cells or human peripheral blood lymphocytes in a direct method nor in a metabolic activation method. Moreover, chromosomal aberrations were not detected in bone marrow cells of CD-1 male mice, even at high rhEPO concentrations (100 000 IU/kg) in vivo. Consequently, it was concluded that errors in the process of enucleation or differentiation of the erythrocytes should be equally considered as possible mechanisms for the increased frequencies of MNPCE and MNRET alongside induction of DNA damage or errors in the process of DNA repair.