Toxicology of Industrial Compounds

V.ROGIERS ET AL. 213
chromatography and reversed phase HPLC, the GST subunits of cocultured
rat hepatocytes were purified and separated (Vandenberghe et al,
1988a). Alterations comparable to those observed for mono-cultures
suggested changes towards a more ‘foetal-like’ state. Less variations,
however, were noticed in the GST subunit pattern of co-cultured
hepatocytes when various media conditions were compared. Incorporation
of 35 S-methionine in the medium showed the ability of co-cultured rat
hepatocytes to synthesize the different GST subunits and suggested that
changes in GST subunit expression under various culture conditions were
the result of in vitro ‘de novo’ synthesis (Vandenberghe et al., 1990a).
Northern blot analysis, using specific cDNA probes showed that the
mRNA levels encoding GST subunits 1/2, 3/4 and 7 were very dependent
on the culture medium.
Again in co-cultures, the changes observed were much less marked than
was the case for mono-cultures (Morel et al., 1989; Vandenberghe et al.,
1990b). As already mentioned for conventionally cultured rat hepatocytes,
phenobarbital had inducing effects on all the GST subunits, but to a
different extent for each subunit (Vandenberghe, 1989). The increased
steady-state mRNA levels observed in co-cultures after phenobarbital
exposure were the result of an increased transcriptional activity of the GST
genes together with a stabilizing effect of the compound (Vandenberghe et
al., 1991).
Also of interest is that the hormonal regulation of GST is maintained in
co-cultures of male rat hepatocytes. 17β-Oestradiol, triiodothyronine and
thyroxine cause a significant decrease in GST activity. Both the overall GST
activity and in particular that of GST 3–3 and 3–4 are decreased (Coecke
et al., 1995c). In contrast, male sex hormones and human growth hormone
had little effect on the overall activity. The effects of triiodothyronine and
thyroxine were particularly oriented towards GST subunits 3 and 4 and
towards an as yet unidentified GST subunit, which was significantly
increased (Coecke et al., 1995c). 17β-Oestradiol shifted the GST subunit
pattern towards the one observed in freshly isolated cells whereas growth
hormone had no specific effect on the individual protein classes (Coecke et
al., 1995c).
These results clearly show a hormonal regulation of GST in co-cultured
rat hepatocytes, although previous work with mono-cultures failed to
prove any direct effect (Gebhardt et al., 1990). The effects are most
pronounced for the Mu-class GSTs. In man, Mu-class GST genes are
structurally very similar to the rat genes and are of particular interest
because 45 per cent of the European population fails to express a
transferase at the GST M 1 locus (Zhong et al., 1993). It is this class of GSTs
that is very effective in deactivating mutagenic and carcinogenic epoxides.
UDP glucuronyltransferases (UDP-GT) have been much less studied in
cocultures than GST. From the work of Niemann et al. (1991) it appears