Environmental and Molecular Mutagenesis - Legacy Tobacco ...

Few are commonly used . Mammalian cell assays hold a special position in this
Irategy because of the hroad spectrum of damage that is theoretically
deteccable with them, but which is not acceasable to prokaryotic assays .
Reasons for this privilege include the higher order structure of mammalian
chromosomes, the different repair mechanisms and the different chemical and
metabolic reactions possible in mammalian as compared with prokaryotic cells .
Fundamental to the acceptability or otherwise of mammalian cell assays is
their success in rodent carcinogen prediction, although they also have value
in biochemistry . Their primary function will be discussed and it will be
concluded that certain assays have reasonable sensitivity (eg ., mouse lymphoma
cell tk locus and sister-chromatid exchange assays), while others have higher
specificity (eg ., hprt locus and primary hepatocyte unscheduled DNA synthesis
assays) . Chromosomal abberition assays show moderate sensitivity and
specificity . These properties of the assays can lead to proposals that some
of them should be discontinued or that, if retained, their use shoul~ be
-d bv scientific and ethical objectives .
365
UV-INDUCED CYTOGENETIC DAMAGE IN WILD-TYPE AND THYMIDINE KINASE DEFICIENT FRIEND MOUSE
ERYTHROLEUKAEMIA CELLS . V .J .McKelvey and P .G .McKenna, Biomedical Sciencee Research
Centre, University of Ulater, Coleraine BT52 1SA, N .Ireland .
Deficiency of the salvage pathway enzyme thymidine kinase (TK) in Friend mouse
erythroleukaemia cells results in increased sensitivity to cell killing and mutagenesis
following UV-irradiation (McKenna,P .G . and Hickey .I . (1981) Cell Biol .Int .Reps . 5 :555) .
Work with other malignant cell lines indicates that TK deficiency only confers eZevated
sensitivity in those cell lines which are normally proficient in DNA excision repair as
evidenced by their ability to undergo unscheduled DNA synthesis (UDS) (McKenna,P .G .,
Yasseen,A,A . and McKelvey,V .J . (1985) Somat .Cell Mo1 .Genet . 11 :239) . Further work has
shown that TK deficiency in Friend cells does not inhibit UDS dccurring (McKenna,P .G .
and McKelvey,V .J . (1986) Somat .Cell Mol .Genet . 12 :325) . In this study vild-type clone
707 Friend cells and two TK deficient eubclonea 707BUE and 707BUF, having TK activities
of 1 .4% and 0 .7% that of wild-type cells respectively, were examined for cytogenetic
aberrations following UV-irradiation . Three doses of UY light were used, namely 2 .4,
4 .8 and 7 .2 J/m and UV-irradiated cultures were harvested for chromosome spreads at 15
hours following treatment . Fifty randomly selected chromosome spreads per treated and
control culture were scored for thirteen types of cytogenetic aberrations . The
frequency of very severely damaged (pulverised) cells was observed to be considerably
greater in each of the two TK deficient aubclones,707BUE and 7078UF, relative to wildtype
clone 707 cells, for each UV treatment examined . Increased UV-aensitivity in the
TK deficient subclones was also reflected in the total aberration frequencies exhibited
by the 3 cell types . The imp rtance of thymidine kinase for accurate DNA repair of UVinduced
damage ia indicated .(The support of the Ulster Cancer Foundation is acknowledge40
366
METABOLISM OF FOOD MUTAGENS WITH PURIFIED AND cDNAl EXPRESSED CSCfOCMROMES P-450 . M+
McManus, W .M . Byrgess, M .E . Ver3nese, J .S . Felton , M .G . Rnize , E .G . Snyderwine ,
L .C . Quattrochi and R .Y . Tukey . Department of Clinical Pharmacology, 2Flinders
University, Australia, Lawre3ce Livermore National Laboratory U .S .A ., National
Institute of Health, U .S .A ., & University of California, San Diego, U .S .A .
We have investigated the specificity of six purified forms of rabbit liver cytochrome
P-450 to activate the food derived haterocyclic amines . IQ and PhIP, to mutagens
in the Ames test . The polcyclic hydrocarbon inducible isozymes Forms 4 and 6
were efficient activators of both these compounds whereas, Forms 2,3b,3c and 5 were
inactive in metabolizing IQ and PhIP to mutagens . The number of revertants produced
in the Ames test per 10 pmol of Form 4 with IQ and PhIP as substrates were 136, 160
and 1,521, respectively . In the presence of 10 pmol of Form 6 and IQ as substrate
16,000 revertants were obtained, whereas PhIP gave 4,577 revertants . When MeIQ, MeIQx
and DiMeIQx were used as substrates Form 4 was at least 3-fold more efficient than
Form 6 in activating these compounds . The gene of the human equivalent of the rabbit
cytochrome P-450 Form 6(P450IA1) and a human cDNA equivalent to Form 4(P450IA2) have
been expressed in Coa-1 cells and their capacity to activate the heterocyclic amines
IQ, PhIP,MeIQ,MeIQx and DiMeIQx was determined . Both the human P4SOIA1 and P450IA2
were capable of activating these amines to mutagens . The order of mutagenicity using
either P450IA1 or P450IA2 expressed cell lysates as the activation source were MeIQx >
IQ > DiMeIQx > MeIQx > PhIP, respectively . The IC50's for a-naphthoflavone inhibition
of the expressed human8P450IA1 and P450IA2 activities with IQ as a substrate in the
Ames test were 2 x 10 and 2 x 10- N, respectively . These data indicate that the
human P4501A subfamily is functionally similar to its rabbit counterpart .
http://legacy.library.ucsf.edu/tid/clb93d00/pdf
1989 EMS Abstracts 127
Notes