Micronucleus assay for mouse alveolar Type II and Clara cells.

Autor: Lindberg HK; New Technologies and Risks, Work Environment Development, Finnish Institute of Occupational Health, FI-00250 Helsinki, Finland., Falck GC, Catalán J, Santonen T, Norppa H
Jazyk: angličtina
Zdroj: Environmental and molecular mutagenesis [Environ Mol Mutagen] 2010 Mar; Vol. 51 (2), pp. 164-72.
DOI: 10.1002/em.20520
Abstrakt: The objective of our study was to develop a micronucleus (MN) assay for detecting genotoxic damage after inhalation exposure in mouse alveolar Type II and Clara cells, potential target cells for lung carcinogens. Ten male C57BL/6J mice were exposed to ethylene oxide (630 mg/m(3)) for 4 hr via inhalation; 10 unexposed mice serving as controls. 72 hr after the exposure, Clara cells and alveolar Type II cells were isolated using two different methods. Method 1 included a 15-min trypsin lavage and a 2-hr incubation of cell suspension. Method 2 involved a 30-min trypsin lavage, Percoll gradient centrifugation, and a 48-hr incubation for cell attachment. Nitro blue tetrazolium (NBT) -staining was applied to distinguish Clara cells. The frequency of micronuclei (MNi) was scored in NBT-negative cells (defined as Type II cells in Method 2) and NBT-positive cells (Clara cells). To detect possible differences between the techniques, MNi in Clara cells were analyzed from samples prepared by both methods. With Method 2, a clear increase in the mean frequency of micronucleated cells was seen in the exposed mice as compared with the controls, for both alveolar Type II and Clara cells. However, no significant increase in MN frequency was seen in Clara cells analyzed from samples prepared by Method 1. Based on our findings, mouse alveolar Type II and Clara cells seem to be suitable for MN analysis in studies aimed at identifying genotoxic lung carcinogens. Both alveolar Type II and Clara cells can be isolated using Method 2.
((c) 2009 Wiley-Liss, Inc.)
Databáze: MEDLINE
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