Micronucleus assays using cytochalasin-blocked MCL-5 cells, a proprietary human cell line expressing five human cytochromes P-450 and microsomal epoxide hydrolase.

Abstract

The MCL-5 cell line is a human lymphoblastoid TK+/- cell line that constitutively expresses a relatively high level of native CYP1A1, four other human cytochromes (CYP1A2, CYP2A6, CYP3A4 and CYP2E1) and microsomal epoxide hydrolase, carried as cDNAs in plasmids. The aim of this study was to evaluate this cell line for its suitability for detecting chromosomal anomalies, employing micronucleus formation in cells blocked at cytokinesis as the indicator of clastogenicity. Results from two laboratories ('ICR' and 'Swansea') using different protocols are reported. In the ICR protocol, aflatoxin B1, sterigmatocystin, benzo[a]pyrene, dibenz[a,h]anthracene, 3-methylcholanthrene, cyclophosphamide, N-nitrosodimethylamine, 2-amino-3,8-dimethylimidazo[4,5-f]-quinoxaline, benzidine, 2-aminofluorene, benzene, tamoxifen and omeprazole were tested and gave positive results. Anthracene, phenanthrene and pyrene were negative. In the Swansea protocol, AHH-1 cells, the parent line which constitutively expresses CYP1A1, but does not contain the genetically engineered human cytochromes or epoxide hydrolase, were tested in parallel with MCL-5 cells. Aflatoxin B1, sterigmatocystin, benzo[a]pyrene, N-nitrosodiethylamine, 2-acetylaminofluorene, benzene, omeprazole and tamoxifen were tested and gave positive results. Of these, only benzo[a]pyrene was equally potent in both cell lines. Assays of tamoxifen and omeprazole included kinetochore staining. Omeprazole, but not tamoxifen, induced a significant level of kinetochore-positive micronuclei. The detection of micronucleus formation in these genetically engineered cells appears to be a rapid, eclectic and sensitive method for screening for genotoxic activity in vitro.