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1.
The effects of waterborne nitrite (3 mg/l NO2) on channel catfish were studied to evaluate changes in hematological parameters and phase I-II biotransformation in liver slices. Nitrite-exposed fish had significantly higher methemoglobin, blood and liver nitrite, and significantly lower pO2 than control fish. Total phase I-mediated metabolism of 7-ethoxycoumarin (EC) was not altered in nitrite-exposed fish compared with control fish (291 +/- 43 and 312 +/- 20 pmol/mg/h, respectively). However, phase II glucuronosyltransferase-mediated metabolism of 7-hydroxycoumarin (HC), both as a phase I metabolite of EC and as a parent substrate, was elevated in nitrite-exposed fish (204 +/- 17 and 1007 +/- 103 pmol/mg/h, respectively) as compared to control fish (149 +/- 14 and 735 +/- 87 pmol/mg/h) (P < 0.05). Sulfotransferase-mediated metabolism of HC (as a metabolite of EC and as a parent substrate) was not notably altered in nitrite-exposed fish (95 +/- 16 and 617 +/- 33 pmol/mg protein/h, respectively) as compared with control fish (118 +/- 24 and 575 +/- 55 pmol/mg/h, respectively). These studies indicate that in vivo nitrite exposure and associated changes in hematological parameters do not appear to affect hepatic phase I EC biotransformation in channel catfish. However, subtle but significant changes in phase II glucuronidation, but not sulfation activity, were observed. The mechanism of these alterations is unclear. However, the data suggest that environmentally realistic concentrations of nitrite may affect the dynamics of conjugative metabolism in exposed fish.  相似文献   

2.
Hepatic microsomes and cytosols of channel catfish (Ictalurus punctatus), rainbow trout (Oncorhynchus mykiss), Atlantic salmon (Salmo salar), red tilapia (Oreochromis sp.), largemouth bass (Micropterus salmoides), striped bass (Morone saxatilis), hybrid striped bass (M. saxatilis × M. crysops), and bluegill (Lepomis macrochuris) (n = 8) were used to study the kinetics of phase I (ECOD, EROD, PROD, BROD) and phase II (UDP-glucuronosyltransferase (UDPGT)-, sulfotransferase (ST)- and glutathione-s-transferase (GST)-mediated) reactions. The best catalytic efficiency for ECOD and GST activities was performed by channel catfish, Atlantic salmon, rainbow trout and tilapia. The highest EROD catalytic efficiency was for Atlantic salmon. None of the species had either PROD or BROD activities. Rainbow trout had very similar UDPGT catalytic efficiency to tilapia, channel catfish, Atlantic salmon, largemouth bass and bluegill. Sulfotransferase conjugation had no significant differences among the species. In summary, tilapia, channel catfish, Atlantic salmon and rainbow trout had the best biotransforming capabilities; striped bass, hybrid striped bass and bluegill were low metabolizers and largemouth bass shared some capabilities with both groups.  相似文献   

3.
The effects of waterborne nitrite (3 mg/l NO2) on channel catfish were studied to evaluate changes in hematological parameters and phase I–II biotransformation in liver slices. Nitrite-exposed fish had significantly higher methemoglobin, blood and liver nitrite, and significantly lower pO2 than control fish. Total phase I-mediated metabolism of 7-ethoxycoumarin (EC) was not altered in nitrite-exposed fish compared with control fish (291±43 and 312±20 pmol/mg/h, respectively). However, phase II glucuronosyltransferase-mediated metabolism of 7-hydroxycoumarin (HC), both as a phase I metabolite of EC and as a parent substrate, was elevated in nitrite-exposed fish (204±17 and 1007±103 pmol/mg/h, respectively) as compared to control fish (149±14 and 735±87 pmol/mg/h) (P<0.05). Sulfotransferase-mediated metabolism of HC (as a metabolite of EC and as a parent substrate) was not notably altered in nitrite-exposed fish (95±16 and 617±33 pmol/mg protein/h, respectively) as compared with control fish (118±24 and 575±55 pmol/mg/h, respectively). These studies indicate that in vivo nitrite exposure and associated changes in hematological parameters do not appear to affect hepatic phase I EC biotransformation in channel catfish. However, subtle but significant changes in phase II glucuronidation, but not sulfation activity, were observed. The mechanism of these alterations is unclear. However, the data suggest that environmentally realistic concentrations of nitrite may affect the dynamics of conjugative metabolism in exposed fish.  相似文献   

4.
Conjugation of phenolic xenobiotics and metabolites through sulfation and glucuronidation is an important biotransformation pathway. Sulfotransferases (SULT) are generally considered non-inducible, while some UDP-glucuronosyltransferase (UGT) isoenzymes are co-induced with cytochrome P450-1A by Ah-receptor ligands. To test these assumptions for two fish species, we measured sulfation and glucuronidation of 9-hydroxybenzo[a]pyrene in 3-methylcholanthrene (3-MC) treated channel catfish (Ictalurus punctatus) and in mummichog (Fundulus heteroclitus) from the creosote contaminated Atlantic Wood site in the Elizabeth River, VA. The results show a significant induction of both UGT and SULT activity in 3-MC treated catfish, linked to the expected induction of EROD activity. In mummichog, significant induction of UGT was measured at the contaminated site over the reference site (King's Creek, VA), as well as extremely low SULT activities at both sites. Western blots, using a polyclonal antibody for catfish phenol-type SULT, confirmed the absence of phenol-type SULT in mummichog. Residual, though slightly inducible, SULT activity may be attributed to other SULT isoforms.  相似文献   

5.
In fish, as well as in mammals, it is well known that the cytochrome P450-dependent oxidative metabolism of xenobiotics can generate DNA-reactive species. Moreover, this metabolism is known to be inducible by several compounds of environmental significance, such as polychlorobiphenyls, polycyclic aromatic hydrocarbons (PAHs) and dioxins. Consequently, we studied the relationship between the degree of induction of the cytochrome P4501A, expressed as that of 7-ethylresorufin O-deethylase (EROD) activity, and the level of DNA-adducts, using the post-labelling assay, in the liver of rainbow trout exposed to benzo(a)pyrene (a representative PAH). The results showed a significant 2- to 4-fold increase in EROD activity 2, 4 and 8 days after treatment, paralleled by an increase in DNA-adduct levels. This work further emphasizes the involvement of cytochrome P4501A in the metabolism of benzo(a)pyrene into genotoxic metabolites in rainbow trout.  相似文献   

6.
The carbamate pesticide, aldicarb, demonstrates significant acute toxicity in fish, and is readily biotransformed by most organisms studied. In fish, both the cytochrome P450 (CYP) and the flavin monooxygenase systems (FMO) are involved in bioactivating aldicarb to aldicarb sulfoxide, which is a more potent inhibitor of acetylcholinesterase (AChE). Channel catfish (Ictalurus punctatus), along with many other fresh water species, do not express FMO and are relatively resistant to the effects of aldicarb. This project examined the toxicity, AChE inhibition, metabolism, and toxicokinetic of aldicarb in channel catfish, and compared these values with an aldicarb-sensitive species, rainbow trout, which expresses FMO. Studies of in vitro and in vivo aldicarb biotransformation in catfish suggest that a low rate of bioactivation (10 times slower Vmax), resulting in less initial conversion to the activated metabolite, aldicarb sulfoxide, may be a contributing factor to resistance of channel catfish to aldicarb toxicity. These data are supported by toxicokinetic and enzyme inhibition studies. This work demonstrates that differences in FMO expression among fish species may have significant influence on toxicity resulting from exposure to some xenobiotics.  相似文献   

7.
In order to examine factors that may contribute to the reported resistance of rainbow trout, Shasta strain, to the well-known hepatocarcinogenic effects of 2-acetylaminofluorene (AAF), the in-vitro and in-vivo metabolism of [14C]AAF in trout has been examined. Trout (compared to rat) liver microsomes metabolized AAF more efficiently, producing 3-fold larger amounts of ring-hydroxylated metabolites (7-hydroxy-AAF and 5-hydroxy-AAF), but 5-fold less N-hydroxy-AAF. Freshly isolated trout hepatocytes extensively metabolized AAF to form the same ring-hydroxylated metabolites and their respective glucuronide and sulfate conjugates. N-OH-AAF (plus its conjugates) and covalently-bound AAF derivatives amounted, respectively, to < 1% and 1.4–1.6% of total metabolites. Liver DNA of trout treated with AAF contained a single AAF-DNA adduct identified as N-(deoxyguanosin-8-yl)-2-aminofluorene (the major persistent AAF-DNA adduct found in rat liver). The level of this adduct (12 attomoles/μg DNA) was about 1000-fold lower than the level of AAF-DNA adduct previously reported in rat liver. The data show that trout liver, compared to rat liver, is considerably less efficient in metabolizing AAF to carcinogenic metabolites, and more efficient in forming nontoxic products, thus possibly explaining, in part, the resistance of trout to AAF-induced hepatocarcinogenesis.  相似文献   

8.
The disruption of chromosome segregation which is detected visually by the anaphase aberration (aa) test suggests that unequal amounts of DNA are distributed to daughter cells and possibly to subsequent cell generations. To investigate this possibility trout cell cultures and trout embryos (blastodisc) were exposed to benzo[a]pyrene (B[a]P) and the nitrosamide, MNNG, respectively. They were then examined by flow cytometry to determine if anaphase damage resulted in unequal DNA distribution to daughter cells. Both B[a]P and MNNG produced a significant increase (P < 0·01) in aa in cultured cells after 48 h exposure. These values returned to normal by 10 days in the absence of the genotoxic agents, except for the highest concentration (0·5 μg/ml MNNG), which showed only a 50% recovery by that time. Likewise, the coefficient of variation (CV) of nuclear DNA content of the cells also rose significantly after treatment and remained elevated for as long as 14 days following exposure. Experiments with rainbow trout embryos also showed a significant increase in both aa and CV following exposure to MNNG. Flow cytometric analysis of DNA content of trout cells and embryos following exposure to mutagens showed an unequal distribution of DNA that was transmissible through several cell generations. These findings indicate that visible anaphase aberrations could predict heritable genetic defects such as those associated with aneuploidy.  相似文献   

9.
Induction of hepatic cytochrome P450-dependent microsomal mono-oxygenase by xenobiotics is a well-established phenomenon in teleost fish. As in laboratory mammals, fish possess multiple forms of cytochrome P450 that display overlapping substrate specificity. One such isoform, CYP1A1, which has been cloned and sequenced from rainbow trout, has been shown to be orthologous to rat CYP1A1 and, as in mammals, is inducible up to several hundred-fold by planar aromatic hydrocarbons, PCBs and dioxins. It has been suggested that induction of CYP1A1 orthologues might provide a sensitive biomonitor for environmental pollution by mixtures of such compounds. In the current study, polyclonal antibodies directed against CYP1A1 purified from rat and trout liver were used to monitor induction of the CYP1A1 orthologue in hepatic microsomes from the fresh water species, the channel catfish (Ictalurus punctatus). Catfish from a local fish farm were induced in the laboratory by three daily injections of 50 mg/kg of the PCB mixture Aroclor 1254 and compared with fish taken from a site in central Arkansas—the Bayou Meto, known to be polluted with dioxin. Hepatic microsomal activities towards ethoxyresorufin (EROD) and pentoxyresorufin (PROD) were measured and Western blot analysis carried out with the two antibodies. EROD was elevated in both the Aroclor-treated fish and in the Bayou Meto fish compared with untreated fish farm controls; smaller but significant increases were observed in PROD. Spearman's rank correlations of 0·74 and 0·89 were observed between EROD and immunoquantified cross-reactivity towards the rat CYP1A1 and trout CYP1A1 antibodies.  相似文献   

10.
The effects of isosafrol (ISF) or β-naphthoflavone (βNF) treatments on cytochrome P450 (P450) levels in rainbow trout liver were investigated using immunochemical and catalytic techniques. The discrepancies in catalytic activities and ELISA quantification of rainbow trout P4501A1 protein levels between ISF- and βNF-treated fish indicate that important differences exist between the responses induced by βNF and ISF treatments in the rainbow trout liver.  相似文献   

11.
cDNA's coding for cytochrome P4501A1 (CYP1A1), phenol UDP-glucuronosyltransferase (UDPGT) and glutathione S-transferase (GST-A) were cloned and sequenced from an expression library prepared from the liver of a 3-methylcholanthrene (3-MC) induced plaice. Plaice CYP1A1 and Phenol UDPGT display a high degree of structural conservation with homologous mammalian isoforms and their mRNAs were shown to be highly induced in liver after 3-MC treatment of fish. Expression of plaice GST-A, which displays closer homology to GSTs from plants and invertebrates than those of mammals, is repressed after 3-MC treatment.  相似文献   

12.
Baseline data for hepatic xenobiotic metabolizing biomarker enzyme activities were obtained for artificially reared tilapia Oreochromis niloticus, and were compared with those of the plaice (Pleuronectes platessa) and rainbow trout (Onchorynchus mykiss). Basal activities exhibited species variations with notably higher CYP1A and phenol UGT activities and lower GST activity in plaice than the freshwater species. Interspecies relationships between gene families determined by immunoblotting and substrate-activity profiles demonstrated the presence of homologous CYP1A and CYP3A enzymes in all three species, alpha class GSTs in plaice and trout, mu and pi class GSTs in trout and theta class GSTs in plaice and tilapia. CYP1A of tilapia was induced by 3-MC or PBO treatment, whilst CYP3A was induced by PCN treatment.  相似文献   

13.
Hepatic levels of the pollutant inducible enzyme, CYP1A, are strongly suppressed in spawning female fish, a phenomenon attributed to high plasma levels of the female sex steroid hormone, estradiol. To evaluate the contribution of estrogen metabolites to estradiol-mediated CYP1A regulation, we treated primary hepatocytes isolated from juvenile rainbow trout (Oncorhynchus mykiss) with vehicle, 17beta-estradiol, or the estrogen metabolite, estriol, alone and in combination with each other and with the potent CYP1A inducer, benzo[a]pyrene (B[a]P). We found dose-dependent suppression of B[a]P-induced CYP1A activity by both steroids relative to controls. At 10(-7) M doses, estradiol and estriol suppressed B[a]P-induced CYP1A activity by 3- and 2-fold, respectively. Although not statistically significant, mean basal CYP1A activity levels were 15- and 13-fold lower in estradiol and estriol treated hepatocytes, respectively, relative to vehicle treated controls. Combining doses of estradiol and estriol failed to produce synergistic suppression of either basal or B[a]P-induced CYP1A activity relative to treatment with either steroid alone. The observed suppression is well below the often strong suppression observed in spawning female fish. We conclude that factors in addition to estradiol and estriol are likely involved in producing sexual dimorphism in CYP1A expression observed in spawning fish.  相似文献   

14.
Purification of cytochromes P-450 from liver microsomes of β-naphthoflavone (BNF)-fed rainbow trout yielded three apparently homogeneous forms. The major form (LM4b)* appears to be a P-448 type of cytochrome. A minor form (LM4a), having properties very similar to LM4b, was also obtained. In addition, a P-450 form (LM2) was isolated, with properties quite different from LM4a or LM4b, including a high rate of aflatoxin B1 (AFB1) metabolism (Williams & Buhler, 1983c). Antibodies to all three forms were obtained from rabbits. The IgGs prepared against LM4a and LM4b both cross-reacted (forming lines of identity) equally well with both antigens on Ouchterlony plates. Rat P-448 cross-reacted (without lines of identity) with both LM4a- and LM4b-IgG. LM4b-IgG was much more effective than LM4a-IgG for inhibition of LM4a or LM4b reconstituted benzo[a]pyrene (BP) hydroxylase, suggesting that these two antibodies recognize different antigenic sites. The LM2-IgG did not cross-react with any of the other rat or trout cytochromes P-450 examined. Levels of LM2 and LM4b in microsomes from untreated, polychlorinated biphenyl (PCB), phenobarbital (PB) or BNF-treated trout were estimated with an immunological technique involving electrophoresis on SDS-PAGE, followed by transfer to nitrocellulose and staining with either LM 2 - or LM4b -IgG. The ratio of in microsomes from PCB- or BNF-treated rainbow trout was much higher than 1, whereas the reverse was true with microsomes from untreated rainbow trout. These results are consistent with previous observations (Vodicnik et al., 1982) that pretreatment with BNF induced the synthesis of a P-448 type cyytochrome, presumably responsible for the great increase in the metabolism and activation of BP seen in these fish. Conversely, pretreatment with PB did not affect the levels of either LM2 or LM4b. This specific immunological technique should make it possible to assay the levels of these P-450 and P-448 isozymes in various strains of rainbow trout and other species of fish. In addition, the effect of age, sex, diet and exposure to P-450 and P-448 inducers could be examined and, perhaps, utilized to predict the relative risk of certain populations to pollutants activated by these different isozymes.  相似文献   

15.
Aquatic organisms are extensively exposed to phthalate esters. We have investigated in trout the effects of diethylhexylphthalate (DEHP) and dibutylphthalate (DBP) on xenobiotic metabolizing enzymes which have been suggested as possible environmental biomarkers. Rainbow trout (Oncorhynchus mykiss) were waterborne exposed to DEHP (1 mg/l) or DBP (0.1 or 1 mg/l) for 72 h. Another group of rainbow trout received daily for 3 days an intraperitoneal injection of 50 mg/kg of DEHP or DBP. Laurate hydroxylation, ethoxyresorufin-o-deethylation, UDP-glucuronyltransferase activity and glutathione-S-transferase activity were measured in liver and extrahepatic tissues. The phthalate esters have been found not to induce these enzymes; in particular, the results do not support the previously described induction of lauric acid hydroxylase in sea bass treated with DEHP [Comp. Biochem. Physiol. B122 (1999) 253.].  相似文献   

16.
Effects on the hepatic cytochrome P450 1A1 system were investigated in juvenile rainbow trout i.p. injected with three different aromatic containing fractions: kerosene, light gas oil or heavy gas oil, originated from distilled North Sea crude oil. Kerosene treatment resulted in no effect on the P450 1A1 system, light gas oil injection caused a weak induction of EROD activities and heavy gas oil treatment resulted in a prominent induction of EROD activities as well as accumulation of CYP1A1 mRNA and P450 1A1 protein levels. The effects of heavy gas oil were compared with effects of β-napthoflavone (β-NF) on the P450 1A1 system. It was obvious that important discrepancies seemed to exist between EROD activities and corresponding CYP1A1 mRNA and P450 1A1 levels in rainbow trout treated with either heavy gas oil or β-NF i.e. heavy gas oil treatment resulted in higher specific EROD activities (EROD/P450 1A1) compared to β-NF. GC-MS analyses revealed that liver and bile from heavy gas oil treated rainbow trout in addition to naphthalene also contained polycyclic aromatic hydrocarbons such as phenanthrenes, anthracene, pyrenes, fluoranthene benz(a)anthracene and chrysene, while none of these compounds were detected in control trout.  相似文献   

17.
In recent years several studies reported the regulation by growth hormone (GH) of the expression of a variety of P450 forms in mammals. However the effect of GH on the xenobiotic-metabolizing enzymes of fish are still unknown. The aim of this work was to investigate the effects of ovine GH—a growth hormone known to be efficient in trout—on the cytochrome P450 level and on aryl hydrocarbon hydroxylase, aminopyrine-N-demethylase, glucuronyl transferase and glutathione transferase activities in trout. The GH-implanted trout (n = 50) each received a single cholesterol pellet containing ovine GH and were compared to control animals (n = 50) receiving a single cholesterol pellet without GH. After 15 days fish were killed and the liver and gills were excised for the measurement of xenobiotic-metabolizing enzyme activities. GH treatment significantly decreased the level of hepatic cytochrome P450 and the activities of cytochrome P450 dependent monooxygenases. In contrast, no significant effect of the treatment was observed on the glutathione transferase and UDP-glucuronyl transferase activities. Moreover, GH treatment had no effect on branchial phase I and phase II enzyme activities. This study provides evidence that GH level significantly affects the expression of several members of the hepatic cytochrome P450 family in trout.  相似文献   

18.
The process of uptake, metabolism and eventual biliary excretion of xenobiotics by the teleost liver affords an opportunity for alteration within at least the following cells: sinusoidal endothelial, perisinoidal fat-storing (vitamin A-containing),1 hepatocytes and biliary epithelial (of preductules, ductules and intrahepatic ducts). Our understanding of the existence and role(s) in physiologic and pathobiologic phenomena of non-hepatocytic populations of cells in teleost liver has been hampered by the use of immersion fixation procedures obviating resolution and analysis of sinusoidal lining cells and by the subjective nature of light and electron micrographs. The purpose of this study was to quantitatively define the rainbow trout (Salmo gairdneri, Richardson) liver. Computer-assisted morphometry2 was conducted on livers of actively spawning 5 year old male and female trout (Wytheville strain).  相似文献   

19.
Expression of flavin-containing monooxygenase(s) (FMO) correlates with salinity exposure in certain species of euryhaline fish, such as the rainbow trout, Oncorhynchus mykiss. The mechanism(s) by which salinity regulates FMO is unclear. Adult rainbow trout were infused through the dorsal aorta with either cortisol or urea. At 500 ng/ml, cortisol caused a significant increase in FMO-catalyzed thiourea oxidase activity in gill and liver microsomes. FMOI expression, however, was significantly increased by the high cortisol dose only in gill microsomes. The levels of TMAO and urea were not altered by cortisol. In the liver, urea infusion caused an increase in hepatic FMO activity. FMO expression and activity correlated with elevated tissue urea levels, but TMAO concentrations were not related. These results indicate that FMO expression and activity may be partially controlled by the osmoregulatory/stress hormone. cortisol, and concentrations of the organic osmolyte, urea, in the rainbow trout.  相似文献   

20.
Branched chain alkylphenols are weak oestrogen mimics which are present in the aquatic environment and have been implicated in the feminisation of fish. This study reports the biotransformation, bioconcentration and tissue distribution of the xenoestrogen 4-tert-octylphenol (t-OP) in juvenile rainbow trout. Fish were exposed for 10 days to a concentration of 4 micrograms/l of [14C] t-OP in a flow-through system and were sampled after 1, 4, 7 and 10 days of exposure. t-OP residues were extracted from all tissues and analysed by radio-high-performance liquid chromatography. After 1 day of exposure radioactive residues were detected in all tissues and reached steady state conditions in the whole fish after 4 days of exposure. The concentration of t-OP residues were highest in bile, followed by faeces, pyloric caeca, liver and intestine. In these tissues the majority of alkylphenol was in the form of two metabolites which were identified by gas chromatography-mass spectroscopy as the glucuronide conjugates of t-OP and t-octylcatechol. t-OP accumulated as the parent compound in fat with a bioconcentration factor (BCF) of 1190, and in brain, muscle, skin, bone, gills, and eye with BCFs of between 100 and 260. This study suggests that exposure to water-borne alkylphenols results in rapid conjugation and elimination of the chemical via the liver/bile route, but that high amounts of the parent xenoestrogen can accumulate in a variety of other fish tissues.  相似文献   

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