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1.
The incidence of hepatoma, epidermal and other forms of cancerous growths in fish is well documented (Halver, 1967; Matsushima & Sugimura, 1976; Dawe et al., 1964). In fish, as in mammals, cancer may be a result of metabolically activated carcinogens. In mammals the primary enzyme system involved in the activation of environmental carcinogens is the cytochrome P-450 linked mixed-function oxidase (MFO). The hepatic microsomes of the species offish studied contain variable levels of cytochrome P-450. Liver microsomes of the trout Salmo trutta lacustris are surprisingly active in metabolising benzo-[a]pyrene (BP) and other compounds preferentially metabolised by polycyclic aromatic hydrocarbon (PAH)-specific cytochrome P-450. This finding may be significant, because it is apparent that the detrimental effects of PAHs require metabolic activation.We have studied the production of reactive intermediates of BP by following their binding to DNA and by measuring the specific nucleotide-BP metabolite complexes by chromatography. Untreated S. trutta liver microsomes catalyse the production of reactive intermediates of BP which bind to nucleotides of DNA at a rate that is 3–4 times higher than that catalysed by control rat liver microsomes. The most prominent DNA-BP metabolite adducts produced by trout liver microsomes are the nucleoside adduct of BP-7, 8-diol-9,10-epoxide and 9-OH-BP-4,5-oxide and other phenol oxides. In contrast to the trout, another fish species, roach (Rutilus rutilus), has extremely low activity. Although the in vitro binding of BP to DNA is not strictly correlated to in vivo binding or biological effects, our results show that reactive intermediates are formed by trout liver and thus the prerequisite for chemical carcinogenesis or mutagenesis is ful filled. This is further supported by the fact that in Ames's test, trout liver preparations produce mutagenic products from promutagens.  相似文献   

2.
Monooxygenase reactions catalyzed by cytochromes P-450 are paramount in the oxidative metabolism of many xenobiotics, determining both the persistence and effects of numerous types of compounds. Immunological probes are proving useful in evaluating the functions of P-450 isozymes in microsomal preparations from many species. The regulation of specific isozymes by endogenous and exogenous factors can also be evaluated with such probes. Here we describe studies on the activities apparently catalyzed by induced P-450 in fish, evaluated with both polyclonal and monoclonal antibodies to cytochrome P-450E, the apparent major β-naphthoflavone(BNF) or methylcholanthrene(MC)-inducible isozyme purified from scup (S. chrysops) liver.  相似文献   

3.
Cytochrome P-450 monooxygenases catalyze the biotransformation of a great variety of foreign, as well as endogenous, lipid-soluble compounds to more water-soluble products. As in mammals, highest concentration of cytochrome P-450 in fish is found in the liver. However, previous studies have indicated that fish kidney contains relatively high cytochrome P-450-mediated activities.1,2 We have therefore prepared and characterized subcellular fractions from the kidney of rainbow trout suitable for studies on cytochrome P-450-dependent reactions. Furthermore, as in the liver, several cytochrome P-450-mediated reactions in the kidney were induced following treatment of the fish with β-naphthoflavone.  相似文献   

4.
These studies were designed to determine the metabolic capability of the microsomal cytochrome(s) P-450 in spiny lobster hepatopancreas, and to determine how chemicals which selectively modify mammalian monooxygenase activity catalyzed by different cytochrome P-450 isozymes affect the spiny lobster cytochrome P-450. We used a washing procedure to concentrate the hepatopancreas microsomal cytochrome P-450 and remove the inhibitors of monooxygenase activity which are normally present in microsomes. The resulting reparation (MI fraction) was used to determine monooxygenase activity towards benzo[a]pyrene, benz-phetamine, 7-ethoxyresorufin in the presence of either cumene hydroperoxide or NADPH and vertebrate liver cytochrome P-450 reductase. Benzphetamine was the best substrate for the lobster cytochrome P-450, whereas 7-ethoxyresorufin was metabolized very slowly. Studies with chemical modifiers showed that the responses of the lobster cytochrome(s) P-450 were not similar to those of any of the well-characterized cytochrome P-450 isozymes purified from mammalian liver.  相似文献   

5.
Monoclonal antibody directed against a major β-naphthoflavone (BNF)-induced form of teleost cytochrome P-450, P-450E (equivalent to P-450c in rat) was used to immunolocalize this enzyme in liver, gill and heart of scup and trout. Liver sections from both species showed P-450E in the cytoplasm of hepatocytes. No regional differences were observed which might indicate zonation of cytochrome P-450E within subpopulations of hepatocytes. Scup exocrine pancreatic cells were only weakly positive. In the gill of both fish, cytochrome P-450E was restricted to the endothelium (pillar cells) of secondary lamellae, where fluorescence appeared as a chain in longitudinal sections through lamellae and as star-shaped clusters in en face views. Sections of ventricular wall in both species revealed P-450E was restricted to endothelium at margins of muscle bands limiting heart ventricular lumen. Localization in the specific cells of these and other organs may be fundamentally important in understanding the role of cytochrome P-450E.  相似文献   

6.
The monooxygenase cytochrome P-450 is a ubiquitous enzyme well known for its role in the detoxication of drugs and xenobiotics and the biosynthesis of the steroid hormones. The toxicity and carcinogenicity of many chemicals is frequently due to the formation of oxygenated reactive intermediates, which are formed by the same enzymes that effect the detoxication of these chemicals, namely the cytochromes P-450. In mammalia the cytochromes P-450 exist as a group of isoenzymes with overlapping substrate affinities, selectively and differentially induced by their specific substrates or inhibitors. As few enzyme assays have been shown to be specific for a particular cytochrome P-450 isoenzyme the characterisation of these isoenzymes has been dependent on their isolation and purification. It is therefore important to know whether one or more isoenzyme of cytochrome P-450 is specifically concerned with the activation of carcinogens and chemicals, rather than their detoxication, and to develop a specific enzymic assay for this activating enzyme.  相似文献   

7.
The hepatic mixed function oxidase system in the fish differs from that in mammals in its responses to the two classic mammalian inducers. The cytochrome P-448-type inducers (polycyclic aromatic hydrocarbons) stimulate monooxygenase activity, but phenobarbital, a P-450-type inducer, does not.1 We have compared the effects of phenobarbital (PB) and polychlorinated biphenyls (PCB) on the turnover of hepatic microsomal hemoproteins in trout (PCB's are P-448- and P-450-type inducers in mammals, which in fish induce only cytochrome P-448). We show here that neither PCB nor PB treatment changed the turnover rate. However, both the rates of synthesis and degradation were much slower than in the rat.  相似文献   

8.
The metabolism of some xenobiotics can lead to the formation of reactive intermediates with mutagenic/carcinogenic properties. With the carcinogenic PAH these have been identified as bay-region diol-epoxides.1 Phenanthrene, a non-carcinogenic, bay-region containing model PAH, is metabolised in vivo by bony fish at the proximate bay-region position, whereas mammals and other marine organisms mainly form the K-region metabolite 9,10-dihydro-9,10-dihydroxy-phenanthrene.2 We wanted to investigate this difference more closely by studying the regiospecificity of phenanthrene metabolism in vitro both with microsomes from differently pretreated cod and with isolated cytochrome P-450 isozymes from BNF-induced cod.3 Secondly, by preparing antibodies to the major isozyme isolated (called cod P-450c), we investigated the immunochemical properties of the variously treated cod liver microsomes.  相似文献   

9.
Activities of the cytochrome P450 monooxygenase (MO) system and GSH-transferase (GST) were monitored about every 2 weeks in sockeye salmon during their sexual migration and ultimate death. Liver cytosol and microsomes were prepared from the tissue of 50 individuals, five males and five females at each collection. The activities observed were not significantly different between the sexes. Substantial changes occurred in cytochrome P450, cytochrome b5 and the monitored activities during the migration. The most dramatic changes occurred at the spawning grounds and were evident even before spawning occurred. The cytochrome P450 and cytochrome b5 contents increased while the MO and GST activities decreased. Pre-spawning MO and GST activities were similar to those reported for other species undergoing sexual maturation. Migrating salmon offer an interesting model for studying the regulation of sex-dependent and terminal forms of cytochromes P450, MO and phase II activities.  相似文献   

10.
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11.
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   

12.
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   

13.
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14.
The degree of induction of hepatic cytochrome P-450 monooxygenases in fish by various chemicals may vary owing to many factors such as sex, sexual maturity, age, season and environmental temperature. In the present investigation the influences of gonadal steroids and water temperature on the inductive response were studied. The data indicate that gonadal steroids and water temperature modulate the response of the cytochrome P-450 system in rainbow trout to PCB and β-naphthoflavone.  相似文献   

15.
In this study we have used cloned gene probes for human glutathione peroxidase (GPX), rat cytochrome P-450IVA1 and rat cytochrome P-450IIE1 to detect homologous sequences in RNA from the hepato-pancreas of Mytilus edulis. The presence of sequences hybridising to the GPX and P-450IVA1 probes, but not to the P-450IIE1 probe, confirms the ancient origin of the former genes and indicates that conserved-sequence DNA probes from higher organisms can be used to examine the structure and function of genes of environmental interest in marine organisms.  相似文献   

16.
The effects of the mammalian inhibitor ellipticine (5,11-dimethyl-[6H]-pyrido[4,3b] carbazole) were examined in a mechanistic study of the cytochrome P450 monooxygenase system of control and β-naphthoflavone (βNF)-induced hepatic microsomes of Platichthys flesus. Ellipticine was indicated to bind to the haem moiety of cytochrome P450s (gave type II binding spectra) and to inhibit the transfer of electrons from both the hydrophobic binding site of cytochrome P450 reductase (P450R) to P450 (inhibited P450R reductase activity) and the hydrophilic binding site of P450R to soluble electron acceptors (inhibited NAD(P)H-cytochrome c reductase activity). No effect was seen on cytochrome b5 reductase activity. Ellipticine inhibition indicated the involvement of (i) P450R (possibly also P450s) in NADPH- but not NADH- dependent hydroxyl radical production, and (ii) electron transfer and P450/P450R interaction in NADPH-dependent cytochrome P450 1A-catalysed monooxygenation (7-ethoxyresorufin O-deethylase activity and benzo(a)pyrene (BaP) metabolism). Differential effects of ellipticine on cumene hydroperoxide (CHP)-dependent BaP metabolism (P450 peroxidase activity) with CHP concentration indicated the existence of at least two forms of P450 with different substrate affinities for CHP, and different mechanisms of formation for protein adducts and free metabolites. Overall, the studies indicate the primary site of action of ellipticine in P. flesus is binding between Fe3+-P450 and P450R.  相似文献   

17.
The hepatic microsomal monooxygenase system of fish-eating sea birds has received little attention considering the vulnerability of this group of animals to environmental pollution. Knight et al.1 measured monooxygenase activities of six species of fish-eating sea birds towards the organochlorine pesticide aldrin and the dieledrin analogue HCE (1,2,3,4,9,9-hexachloro-1,4,4a,5,6,7,8,8a-octohydro-exo-7,8-epoxy-1,4-methanonaphaline) and found very low activities in all species except the puffin (Fratercula arctica), activities more comparable to those of the fish than those of terrestrial birds. The aims of the present study were to compare the aldrin epoxidase and HCE hydroxylase systems of the razorbill (Alca torda) and the puffin with those of the male rat. Kinetics of the two reactions were compared at substrate concentrations down to environmentally realistic levels and the forms of cytochrome P-450 of the birds were partially characterised and compared with forms equivalently from untreated male rat liver.  相似文献   

18.
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   

19.
Bivalve and gastropod molluscs readily accumulate polynuclear aromatic (PNAH) and other hydrocarbons from the environment and are widely used in environmental monitoring programmes.1 The response of the cytochrome P-450 monooxygenase or mixed function oxidase (MFO) system to organic xenobiotics is of interest from the comparative viewpoint and in biological effects monitoring.2 We have studied the bivalves Mytilus edulis (mussel) and Cardium edule (cockle) and the gastropod Littorina littorea (periwinkle) exposed to hydrocarbons, experimentally or naturally in the field. The general trend of response in digestive gland microsomes was an increase in cytochrome P-450 content and NADPH-cytochrome c reductase (NADPH-CYTCRED) activity but no increase in benzo[a]pyrene hydrolyase (BPH) activity. Sex and seasonal interactions were evident. We conclude that aspects of the responses may be peculiar to the Mollusc and that NADPH-CTYCRED possibly offers potential for this phylogenetic group as a specific indicator of biological impact by organic pollution.  相似文献   

20.
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   

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