Page 711 - The Toxicology of Fishes

Biomarkers 691

al., 1998; Sanders, 1990, 1993). While HSP70s can be found in the cytoplasm and nucleolus, HSP60
(chaparonin) is a mitochondria protein (Schlesinger, 1994). Each is highly conserved, and antibody as
well as nucleotide probes to mammalian forms have been shown to recognize homologous forms in
numerous fish and invertebrate species (Dyer et al., 1991, 1993; Iwama et al., 1998; Sanders, 1993;
Stegeman et al., 1992). The cytosolic HSP90 protein is an integral part of several receptor complexes
(i.e., Ah and steroid receptors) and is induced in catfish, medaka, and fathead minnows following exposure
to detergents and PAHs (Villalobos et al., 1996). Other studies observed induction in rainbow trout
following arsenite exposure (Kothary and Candido, 1982). Unlike the other HSPs discussed in this
section, HSP30 actually serves an enzymatic function known as heme oxygenase, which catabolizes
prosthetic groups to biliverdin, which is then converted to bilirubin by biliverdin reductase (Stegeman
et al., 1992). Induction of HSP30 protein and activity as a result of chemical exposure have been observed
in only relatively few fish species (Ariyoshi et al., 1990; Brown et al., 1993; Grosvik and Goksøyr, 1996;
Kothary and Candido, 1982; Sanders, 1993; Schlenk et al., 1996a). In most of these studies, consistent
induction was observed following exposure to cadmium or arsenite; however, exposure to phenylhydra-
zine and high lipid diets appears to increase concentrations of HSP30 in Atlantic salmon (Lunde et al.,
1998). Because HSP30 is involved in catabolism, induction of HSP30 has been shown to be inversely
proportional to CYP1A and total CYP activity (Schlenk et al., 1996b; Stegeman et al., 1992); thus,
HSP30 may prove to be a confirmatory marker of CYP repression.
With the exception of the HSP70 family of proteins, little work has been done to calibrate or characterize
these proteins in fish; hence, caution should be used in evaluating data using these proteins alone without
other better characterized systems (Kohler et al., 2001). In laboratory studies with rainbow trout, HSP70
levels in juveniles were significantly increased in gills of juveniles exposed to a mixture of cadmium,
copper, lead, and zinc for 21 days, in both water and food (Williams et al., 1996). This response correlated
with decreases in whole-body potassium and increased concentrations of whole-body metals in juveniles
(Farag et al., 1994). Interestingly, no relationship was observed in livers of juveniles or any tissues of
adults (Williams et al., 1996). Evidence supporting HSP70 as a biomarker of effect was also observed
in β-naphthoflavone (BNF)-treated rainbow trout, in which altered metabolic status of the liver as
evidenced by lower phosphoenolpyruvate carboxykinase (PEPCK), lactate dehydrogenase, and
3-hydroxy-acyl-coA dehydrogenase activities correlated to hepatic HSP70 expression. This study also
showed that HSP70 in this species was not modified by handling stress (Vijayan et al., 1997). Relation-
ships between HSP70 expression and ovarian follicular apoptosis were observed in white sucker exposed
to bleached kraft pulp mill effluent (Janz et al., 1997). Seasonal differences in HSP70 expression have
also been observed in wild fathead minnows, Atlantic salmon, yellow bullhead catfish, and rock bass,
with highest levels of expression being in winter and lowest in summer and fall (Fader et al., 1994).
Compelling evidence suggests that HSP70 may be a useful bioindicator of general cellular stress
relating to proteolysis. Although the induced synthesis of the protein is transient, the turnover is much
less rapid and the proteins tend to accumulate upon continued cellular stress (Sanders, 1990). In addition,
the kinetics of induction appear to be longer following chemical-induced stress compared to heat-induced
stress, and recovery is not achieved until several days following exposure to metal, presumably due to
accumulation of the metal in the cell (Stegeman et al., 1992). Overall, significant gaps remain in
characterizing basal activities of piscine HSPs, especially regarding potential susceptibility, gender-
related, and developmental differences. Thus, it is suggested that these proteins be used in conjunction
with other acute phase protein markers (i.e., MT) to verify effect.

Multidrug Resistance (P-Glycoprotein)
P-Glycoprotein is a multi-substrate membrane transport protein involved in the resistance of tumor cells
to chemotherapy (Juliano and Ling, 1976). Also known as the multidrug resistance (MDR) or multix-
enobiotic resistance (MXR) mechanism, P-glycoprotein expression has been shown in numerous studies
with invertebrates and humans to be regulated by exposure to various classes of organic and inorganic
chemicals (Kurelec, 1992, 1997). Because of this ability to be upregulated in response to multiple
chemicals, MDR would not be considered to hold promise as a biomarker of exposure to a specific class
of chemicals; however, increased expression has been observed, especially in neoplastic tissues, and