Gr. Douglas et al., TOWARD AN UNDERSTANDING OF THE USE OF TRANSGENIC MICE FOR THE DETECTION OF GENE-MUTATIONS IN GERM-CELLS, Mutation research. Genetic toxicology and environmental mutagenesis, 388(2-3), 1997, pp. 197-212
Recently-developed transgenic models have provided unprecedented acces
s to rodent somatic and germ line tissues for the study of gene mutati
on in vivo. While mutations in germ cells are considered an important
aspect of any regulatory assessment of the risks posed by chemicals, c
urrently-available conventional tests, which involve the study of thou
sands of offspring make it impractical to test large numbers of chemic
als, for the induction of inherited gene mutations. When effects in ge
rm cells per se, rather than offspring are acceptable targets, transge
nic mouse assays may provide a practical alternative. As part of an in
ternational collaborative study to begin to determine the reliability,
efficacy, and role of such assays, lacZ transgenic mice (Muta(TM) Mou
se) were treated with single i.p. doses of ethylnitrosourea (ENU), met
hyl methanesulfonate (MMS), and isopropyl methanesulfonate (iPMS), and
mutant frequencies determined using phenyl-beta-D-galactoside (p-gal)
positive selection. For studies using germ cells, the selection of sa
mpling times and target cells is crucial. Spermatagonial stem cells an
d cells in post-spermatagonial stem cell stages are the critical targe
t cell populations of regulatory importance. Cell populations within t
hese categories were studied by sampling germ cells isolated from semi
niferous tubules and spermatozoa from the epididymis at 91 days and 25
days after treatment. The data show that ENU and iPMS induced mutatio
ns in post-spermatagonial stem cells and spermatagonial stem cells. Ho
wever, MMS did not induce mutations in either cell type, or at either
sampling time, at doses approaching lethality. This result is possibly
because MMS induces preferentially large lesions and chromosomal aber
rations (as opposed to point mutations), which are not readily detecta
ble with bacteriophage-based shuttle vectors. Since MMS-induced specif
ic locus and dominant lethal mutations are induced only after the mid-
spermatid stage, it is also possible that the timing used missed this
effect. While the ENU and iPMS data in this study demonstrate the suit
ability of the lacZ male transgenic mice for the study of gene mutatio
ns in post-spermatagonial stem cells and spermatagonial stem cells by
sampling cells isolated from seminiferous tubules at selected times af
ter treatment, the MMS results do not answer fully whether transgenic
mouse mutation assays can detect mutations resulting from lesions indu
ced after the mid-spermatid stage when most cellular processing is ret
arded. Nevertheless, it appears clear from presently available informa
tion, that the bacteriophage-based lacZ transgenic model is suitable f
or the detection of gene mutations in spermatogonial stem cells, sperm
atocytes, and early spermatids.