Page 208 - The Toxicology of Fishes

188 The Toxicology of Fishes

antisera, indicating common structural epitopes (Clarke et al., 1992a). From purification studies, at least
five UGTs have been identified in plaice liver (George, 1994), and the phenol-conjugating isoform that
was purified to homogeneity displayed a very high activity toward planar phenols and no measurable
activity toward bilirubin or steroids. The bilirubin-conjugating isoform appeared to exhibit both bilirubin-
and phenol-conjugating activity as found in mammals. Reviews of earlier studies on glucuronidation in
fish can be found in Clarke et al. (1991) and George (1994). The picture that emerges from these reviews
is that UGT isoforms are found in a wide variety of fish species and that specific enzymatic activities
are found for bilirubin, steroid and thyroid hormones, and phenolic compounds. As in mammalian
systems, a diverse group of xenobiotic compounds has been identified as substrates for UGTs, among
which are chlorinated phenols, aromatic hydrocarbon metabolites, phthalates, aflatoxin, pesticides, and
antibiotics. Comparison of UGT activities among species was found to be a problematic task primarily
due to the non-optimization of detergent concentrations in assays in most published studies. The enzymes
also display maximal activity at 37°C (Clarke et al., 1992b); thus, corrections for temperature should
be used for intercomparisons.

Tissue Distribution
In fish species, UGT activity is usually high in liver and intestine, but measurable activities have also
been found in gill, kidney, and muscle tissue (Clarke et al., 1991; George et al., 1998; James et al., 1998;
Singh et al., 1996). UGTs play an important role in gonadal tissues; in addition to the regulation of
steroid hormones, UGTs play a role in the production of sex pheromones in fish (Lambert and Resink,
1991). Testosterone UGT activity is present in liver, testis, and intestine (Clarke et al., 1992b), and
glucuronidation of pregnenolone and androstenedione has been demonstrated in vitro with testicular
preparations (Andersson, 1992). Glucuronidation of bilirubin is confined to liver in both plaice and
salmon.

Regulation of UGTs
Coregulation of both CYPs and UGTs occurs in mammals, and prototypical inducers such as clofibrate,
PAHs, phenobarbital, and pregnenolone-16α-carbonitrile (PCN) differentially induce expression of both
CYP and UGT isoforms. The degree of upregulation of UGT activity is generally some two- to threefold.
UGT1A1 (bilirubin conjugation) is induced by the hyperlipidemic agent (and peroxisomal proliferator)
clofibrate. In common with a number of phase I and II genes, UGT1A6 (planar phenol conjugation) is
induced by interaction of the Ah receptor with an XRE in its promoter region. This coregulation of
CYP1A and UGT1A6 in mammals and CYP1A and UGT1B1 in fish has been shown to facilitate
detoxification of PAHs such as BaP. Mammalian CYP2B and CYP3A and UGT1A genes are also induced
via a nuclear pregnane X receptor and a constitutive androstane receptor. Interestingly, these recognition
motifs are also present in the zebrafish UGT6220 gene (George, unpublished data). Mammalian steroid
UGTs are induced by PCN. Several reports indicate a modest induction of phenol UGT activity in fish
from polluted environments and after experimental PAH exposure. To study the induction of AhR-
activated enzymes, two PAHs are often used: 3-methylcholanthrene (3-MC) and β-naphthoflavone (BNF).
Both compounds have been used as CYP1A and phenol UGT inducers in a number of fish species (Table
4.9). Maximal induction of CYP1A is usually found around 3 days after single treatment, but phenol
UGT induction appears to be slower, with a maximum induction occurring around 8 days after treatment.
In general, induction of EROD activity can be up to 250-fold, but UGT activity is never induced more
than 3-to 6-fold. Two species, cod (Gadus morhua) and gilthead sea bream (Sparus aurata), displayed
little or no response to AhR ligands as phenol UGT inducers (Goksøyr et al., 1987; Pretti et al., 2001).
It must be noted, however, that multiple isoforms in mammals, including the constitutive steroid isoforms,
also conjugate 4-nitrophenol. Immunoblot and northern blot analyses of xenobiotic-treated plaice have
shown that induction of UGTs appears to be tissue specific (Clarke et al., 1992a). Treatment with the
PAH (3-MC) increased phenol-conjugating activity and a 56-kDa immunoreactive peptide in liver by
approximately 1.7-fold. The bifunctional PAH-type inducer BNF caused an induction (approximately
three- to fourfold) of UGT1B1 mRNA only in intestine (Leaver et al., unpublished data). Although
clofibrate did not appear to induce phenol- or bilirubin-conjugating activities (Clarke et al., 1992a),