Page 381 - The Toxicology of Fishes

Liver Toxicity 361

which species have evolved over a long enough period of geological time have members that form C27
bile alcohol sulfates, species that are in the process of transition (selecting one of the three pathways),
and species that form C24 taurine-conjugated acids. Hence, bile acids in vertebrates can be broken down
into three classes based on side-chain functional group: C27 bile alcohols and C27 and C24 bile acids.
Bile alcohols, which occur in many fish species, are conjugated by esterification of the terminal C27
hydroxy group with sulfate, whereas bile acids are typically conjugated by N-acyl amidation of the
terminal C27 or C24 carboxyl group with taurine or glycine (Moschetta et al., 2005).
In terms of bile synthesis and composition, the majority of fish fall into two groups: ancient (utilizing
C27 bile alcohol sulfates) and modern (utilizing C24 taurine-conjugated acids). C27 bile alcohol sulfates,
characteristic of a more primitive bile synthetic pathway, are ubiquitous among vertebrates. They are
the dominant bile salts of ancient mammals (e.g., elephant, manatee, rhinoceros) and the major bile
constituents in cartilaginous fish, herbivorous bony fish, and some amphibians. In the few fish species
examined, the ratios of bile acids to bile alcohols differ from those of mammals; for example, medaka
synthesize bile using the classical pathway (a transition fish), with approximately 50% C27 taurine-
conjugated acids and 25% C24 taurine-conjugated acids, while a sulfated bile alcohol, scymnol sulfate
(ScyS), is the major bile salt in the little skate (Raja erinacea), and 5-cyprinol sulfate, a more common
piscine bile alcohol, is the major bile salt in zebrafish. Similar differences in bile acid/bile alcohol
composition are described in the coelacanth (Latimeria chalumnae) and Japanese eel (Kihira et al., 1984;
Masui et al., 1967). Of interest is that bile alcohols are known to be stable at 4°C, which may be of
evolutionary significance in cold-water species.

Bile Transport
The vast majority of the bile salt pool is localized to enterohepatic circulation and is regulated in mammals
by a suite of distinct transmembrane transporters in hepatocytes, BECs, and enterocytes (Trauner and
Boyer, 2003). Bile salt homeostasis is also governed through tight negative feedback control via tran-
scriptional regulation; for example, after intestinal absorption bile salts return to liver where they regulate
their own synthesis (see discussion on nuclear regulators). Hepatobiliary transport mechanisms deter-
mining uptake and excretion of bile salts and other biliary constituents in liver and extrahepatic tissues
are well characterized in mammals and are likely well conserved across vertebrate species (with antic-
ipated subtle species differences). Transmembrane transport proteins, localized to both sinusoidal and
apical membranes of hepatocytes and biliary epithelia, have been functionally characterized, cloned, and
studied in transgenic animal models, and several genes regulating transporter synthesis have been
identified through mutations in inherited forms of cholestasis (Boyer, 1996; Pauli-Magnus and Meier,
2005; Trauner and Boyer, 2003).
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The primary mammalian hepatocytic transporters include the sinusoidal Na taurocholate cotransport-
ing polypeptide (NTCP; SLC10A1), organic anion-transporting polypeptides (OATPs; SLC21A), the
organic cation family of transporters (OCTs), multidrug resistance-associated proteins (MRP1–6;
ABCC1–6), and members of the family of multidrug resistance P-glycoproteins, such as MDR1 (ABCB1),
MDR3 (ABCB4), and the apical bile salt export pump (BSEP) (ABCB11) (Boyer, 1996; Groothuis and
Meijer, 1996; Pauli-Magnus and Meier 2005; Sturm et al., 2001b; Trauner and Boyer 2003). Bile salt
transporters have also been identified in biliary epithelia, enterocytes, the renal proximal tubule of the
kidney, and placenta. An important characteristic of MDR-encoded gene products (e.g., MDR1, MDR3)
is their broad specificity; they are able to transport an array of structurally and functionally diverse
substrates, including anticancer drugs such as paclitaxel, vinca alkaloids, anthracyclines, and epipodo-
phyllotoxins (Ueda et al., 1997), as well as numerous other classes of compounds such as calcium
channel blockers, HIV protease inhibitors, hormones, pesticides, cyclic and linear peptides, and immu-
nosuppressive agents (Stouch and Gudmundsson, 2002).
Mammalian bile salt transport occurs via both Na -dependent and Na -independent mechanisms. The
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primary Na -dependent transporter is the sinusoidal NTCP (Ntcp in rodents), which is exclusively
expressed in hepatocytes and mediates the hepatocellular uptake of bile salts (Fardel et al., 2002;
Groothuis and Meijer, 1996; Lecureur et al., 2000). In addition to bile salts, rat Nctp has been shown
to support the hepatic uptake of estrone-3-sulfate, dehydroepiandrosterone sulfate, and the thyroid