Aryl Hydrocarbon Hydroxylase Levels Research Articles

Role of Prosopis cineraria against N-nitrosodiethylamine-induced liver tumor in rats with reference to marker enzymes and nucleic acid contents

The effect of methanol extract of Prosopis cineraria against experimental liver tumor in rats was studied. Liver tumor was induced by the administration of N-nitrosodiethylamine (200 mg/kg) and it was promoted by phenobarbital administration. Methanol extract (200 and 400 mg/kg) was administered to determine the protective activity. Administration of methanol extract suppressed the liver tumor effectively as revealed by the decrease in elevated levels of aryl hydrocarbon hydroxylase, lactate dehydrogenase, g-glutamyl transpeptidase (g-GTP), 5’nucleotidase, deoxyribonucleic acid (DNA) and ribonucleic acid (RNA). We found that methanol extract may extend its protective role by modifying the levels of marker enzymes and nucleic acid contents.

AHH activity, tissue dose and DNA damage in different tissues of the scallop Chlamys farreri exposed to benzo[ a]pyrene

A collaborative study was performed on scallops ( Chlamys farreri) exposed to 0.5, 3 and 10 μg/L benzo[ a]pyrene (B[ a]P) for 20 days. The levels of aryl hydrocarbon hydroxylase (AHH) activity, B[ a]P accumulation and DNA strand break were assayed in the gill and digestive gland. Results showed that AHH activity and B[ a]P accumulation were significantly related to B[ a]P dose. AHH activity was induced and then became stable gradually. B[ a]P accumulation increased first and showed an incoming plateau. DNA strand break levels in the 0.5 and 3 μg/L B[ a]P groups remained high and significantly different from control values until day 6, followed by a reduction in the gill and no recovery in the digestive gland. The 10 μg/L B[ a]P groups remained significantly lower than control until the end. These results suggested that the application of comprehensive indices may give information on the actual exposure of organisms to pollutants, and also information on toxic effects.

In vitro induction of CYP1A1‐associated activities in human and rodent cell lines by commercial and tissue‐extracted halogenated aromatic hydrocarbons

AbstractDespite reports of species‐specific differences, the rat cell line H4IIE continues to be used to assess the toxicity of many halogenated aromatic hydrocarbon (HAH) contaminants for diverse populations using an approach that relates aryl hydrocarbon (Ah) receptor‐mediated responses by HAHs to that of 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD). To assess the relevance of H4IIE for human populations, we examined levels of aryl hydrocarbon hydroxylase (AHH) and ethyoxyresorufin O‐deethylase (EROD) activities in H4IIE rat liver, HepG2 human liver, and MCF‐7 human breast cells treated with TCDD and three coplanar polychlorinated biphenyl (PCB) congeners (77, 126, 169), alone or in combination, and to fractions extracted from the muscle tissue of Lake Ontario rainbow trout (Oncorhynchus mykiss). H4IIE was generally more sensitive to all HAH treatments. Both AHH and EROD activities were induced in all cells treated with TCDD and PCB‐126, alone or combined with each other. In contrast, neither activity was induced in human cells treated with PCB‐77 or PCB‐169. For inducing concentrations of PCBs, the induction response in all cell lines treated with 5 nM TCDD‐PCB mixtures was generally nonadditive; only the response in H4IIE treated with 10−3 nM TCDD–PCB mixtures was generally additive. With tissue extracts, induction was restricted to H4IIE cells treated with TCDD/PCB‐or organochloride pesticide‐containing fractions. Our initial results suggest that differences in the properties of each HAH or in levels of Ah receptor in each cell line are not likely to underlie the significant differences in the murine/human induction responses.

Modulatory influence of alcoholic extract ofOcimumleaves on carcinogen‐metabolizing enzyme activities and reduced glutathione levels in mouse

The present study reports the modulatory influence of alcoholic extract from the leaves of Ocimum sanctum on the activities of cytochrome p-450, cytochrome b5, and aryl hydrocarbon hydroxylase enzymes in the liver and glutathione-S-transferase and reduced glutathione level in the liver, lung, and stomach of the mouse. Oral treatment with the leaf extract at 400 and 800 mg/kg body wt for 15 days would significantly elevate the activities of cytochrome p-450 (p < 0.05), cytochrome b5 (p < 0.01, p < 0.001), aryl hydrocarbon hydroxylase (p < 0.05), and glutathione S-transferase (p < 0.05, p < 0.01), all of which are important in the detoxification of carcinogens as well as mutagens. Moreover treatment with 400 and 800 mg/kg body wt of Ocimum extract for 15 days also significantly elevated extrahepatic glutathione-S-transferase (p < 0.05, p < 0.01). The reduced glutathione level was also elevated by treatment with the leaf extract in liver, lung, and stomach tissues (p < 0.01, p < 0.001). Mice fed a diet containing 0.75% butylated hydroxyanisole (positive control) revealed no alteration in the basal hepatic cytochrome p-450 and aryl hydrocarbon hydroxylase level, but hepatic cytochrome b5 and glutathione S-transferase activity in hepatic and extrahepatic organs were elevated in a time-responsive manner (p < 0.05, p < 0.001). The observations suggest further exploitation of the Ocimum leaf extract or its active principle(s) for the chemoprevention of chemical carcinogenesis in different animal model systems.

Observations on vitamin K deficiency in the fetus and newborn: has nature made a mistake?

The microsomal mixed function oxidase system metabolizes xenobiotics (Phase I) to products that, if not activated and conjugated for excretion (Phase II), are capable of forming conjugates with cellular macromolecules, including DNA, resulting in toxic, mutagenic, or carcinogenic events. Benzo(a)pyrene (BP), a polycyclic aromatic hydrocarbon, is a model carcinogen for this system. Vitamin K1 (phylloquinone) is a regulator of BP metabolism. These studies demonstrate that K1 is capable of increasing Phase I metabolism and decreasing glutathione transferase activity (Phase II) in chick embryo liver; that deprivation of K1 reduces BP/DNA adducts in mouse liver and reduces tumor formation in mice given intraperitoneal BP; and that K1 supplementation increases BP induced tumor formation in mice. However, epidemiologic studies indicate that children of mothers who smoke during pregnancy may not be at increased risk of cancer. It is known that the placentas from these pregnancies exhibit markedly increased levels of arylhydrocarbon hydroxylase induced by the polycyclic aromatic hydrocarbons in tobacco smoke, but there is no corresponding increase in this enzyme activity in the fetus in such pregnancies. We suggest that the low vitamin K level is a secondary protective mechanism for xenobiotics, such as BP, that may escape the primary placental screen. The recently described role of vitamin K-dependent Gla protein as ligands for receptor tyrosine kinases, also establishes K as a link in cell growth and transformation. It is proposed that the small total body pool of K1 in the adult, which is sufficient only to meet continuing needs, and the even smaller pool in the fetus are protective. This protective effect of low K1 levels is particularly important in the presence of the high mitotic rates and rapid cell turnover in the avian embryo and mammalian fetus.

Effect of storage conditions of white sucker liver on some MFO activity and metallothionein concentration

The stability of hepatic mixed-function oxygenase (MFO) activity and metallothionein (MT) level in white sucker ( Catostomus commersoni) was investigated in order to develop standardized freezing and storage procedures for liver samples. Feral white suckers were collected in the St. Lawrence River (Québec, Canada). Liver samples were divided into three groups (A, B and C). They were frozen in liquid nitrogen and then stored at −145°C as (A) S10 fraction; (B) homogenates; (C) pieces of liver. Aryl hydrocarbon hydroxylase (AHH) and ethoxyresorufin- O-deethylase (EROD) activities, and MT concentrations were measured on the S10 fraction in all three groups after storage periods of 2, 7, 14, 21, 84 and 168 days. Results showed that EROD and AHH activities and MT concentrations were stable for up to 168 days, regardless of sample preparation. No significant differences were observed for AHH and EROD levels between the three treatment methods. Treatment A (storage of S10 fraction) maintained significantly higher levels of MT than either treatment B or C.

Impact of Quercetin Consumption on Phase-I and Phase-II Drug-Metabolizing Enzymes in Mice.

In the present study Swiss NMRI mice were fed quercetin, a naturally-occurring dietary polyphenolic flavonoid, through their drinking water at dose levels of 0, 1, 3, and 9μg/ml for 8 weeks. There was no change in the hepatic and pulmonary levels of arylhydrocarbon hydroxylase, cytochrome P-450, cytochrome b5, cytochrome c-reductase, or UDP-glucuronyltransferase due to quercetin feeding. However, hepatic activity of glutathione-S-transferase was significantly increased at 3 and 9μg/ml dose levels; and that of the pulmonary enzyme, at 9μg/ml dose. Levels of reduced glutathione were significantly elevated in liver and lungs. These increases in glutathione-S-transferase and reduced glutathione may protect the liver and lungs from the carcinogenic insult of chemicals through the detoxification process.

Xenobiotic-metabolizing enzymes and benzo[ a]pyrene metabolism in the benzo[ a]pyrene-sensitive mutant strain of Drosophila simulans

Basal levels of aryl hydrocarbon hydroxylase, epoxide hydrolase and glutathione S-transferase enzyme activities, cytochrome P-450 content and inducibility of enzymes with phenobarbital were found to be similar in the microsomes of D. simulans mutant strain 364 yv, which is sensitive to the toxic and mutagenic effects of benzo[ a]pyrene (BP), and of the wild resistant Turku strain. In contrast, increases in the rate of BP turnover per molecule of cytochrome P-450, intensity of the hemoprotein band with apparent molecular weight 56,000 and the yield of BP 7,8-dihydrodiol and 9,10-dihydrodiol occurred only in microsomes of BP-pretreated 364 yv flies but not of Turku ones. It is likely that BP induces an aberrant form of cytochrome P-450 in 364 yv flies with a rare mutation in one of the P-450 regulating genes.

Monocyte aryl hydrocarbon hydroxylase (AHH) activity mimics Kupffer cell and hepatocyte AHH activity in an animal model of liver disease

We have previously reported that monocyte aryl hydrocarbon hydroxylase (AHH) activity is depressed in patients with liver disease and is decreased more in cirrhosis than in early stage liver disease. To determine if monocyte AHH activity reflects liver AHH activity, we studied an animal model of cirrhosis, i.e., yellow phosphorus induced cirrhosis in the pig. AHH activity was detectable in monocytes isolated from peripheral blood of normal pigs (0.32 +/- 0.13 nmol.mg-1 P.h-1, n = 11) and was comparable to the level of AHH activity in hepatic Kupffer cells isolated from wedge or needle biopsies of livers of normal pigs (0.38 +/- 0.21, n = 7). The AHH level in pig Kupffer cells was approximately 10% of the AHH level in hepatocytes and microsomes. To induce liver disease, pigs were administered yellow phosphorus (0.6 mg/kg) 5 days per week for 16 weeks. At 4 weeks of treatment, monocyte AHH activity was not different from control and liver histology was normal. Depression of monocyte AHH activity was evident at 8 weeks of treatment when liver fibrosis was seen histologically. At 12 weeks of treatment when histology revealed extensive liver fibrosis and collagen levels were elevated, the level of monocyte AHH activity was decreased 67% compared with controls. Similar changes were observed at 12 weeks in Kupffer cell AHH activity (86% decrease) and hepatocyte AHH activity (70% decrease) compared with controls. These results suggest that monocyte AHH activity reflects liver AHH activity and may be a good indicator of change in liver enzyme function in liver disease in the pig model of cirrhosis.

Modulatory influences of clove ( Caryophyllus aromaticus, L) on hepatic detoxification systems and bone marrow genotoxicity in male Swiss albino mice

Modulatory effects observed due to clove administration (0.5%, 1% and 2% w/w in the diet) to Swiss albino mice for 10, 20 and 30 days in the hepatic levels of cytochrome P-450 (Cyt. P-450), cytochrome b 5 (Cyt. b 5), aryl hydrocarbon hydroxylase (AHH), glutathione S-transferase (GST), DT-diaphorase (DTD), acid soluble sulfhydryl (SH) content and radiation-induced malondialdehyde (MDA) formation were recorded. Enhanced GST, Cyt. b 5 and SH levels were observed in all the treatment groups, excepting those maintained on a 0.5% diet for 10 days which did not show significant increase in the GST and SH levels as compared to their respective controls. Significant reduction in Cyt. P-450 and MDA levels was observed in all groups at 30 days duration. While AHH levels remained unaltered by clove administration, DTD activity was elevated by 1% and 2% clove diets at 30 days duration. An in vivo bone marrow micronucleus assay demonstrated that administration of 0.5% and 2% clove diets for 10 days neither significantly induced micronuclei nor could effectively modulate the 7, 12-dimethylbenz[ a]anthracene genotoxicity in mice. The results suggest whole cloves as potential chemopreventive agents.

Co-cultures of hepatocytes with epithelial-like cell lines: Expression of drug-biotransformation activities by hepatocytes

To improve long-term expression of drug biotransformation activities in hepatocytes, we have examined the suitability of several epithelial-like cell lines (MDCK, MS and L-132) for supporting functional co-cultures with rat hepatocytes. Cells were selected on the basis of their compatibility with hepatocytes, formation of stable monolayers in the absence of serum and lack of drug biotransformation activities. The expression of individual elements of the biotransformation system was evaluated in these co-cultures. Co-cultured hepatocytes remained viable and showed a characteristic polygonal shape for more than a week. Depending on the cell line used, levels of aryl hydrocarbon hydroxylase and 7-ethoxycoumarin O-deethylase activities of co-cultured hepatocytes oscillated between 24-47% of their initial value after 4 days in culture. The highest levels of monooxygenase activity were found in hepatocytes co-cultured with MS cells (41-47%). In contrast, these activities decreased to 6% when hepatocytes were maintained in pure culture for the same period. The activities of the conjugating enzymes UDP-glucuronyltransferase and glutathione S-transferase were maintained at nearly the initial levels during the complete period of study, both in pure and mixed-cultures, regardless of the cell line used. MS cells adapted themselves much better to serum-free culture conditions, and the co-culture with rat hepatocyte was technically easier. After one week, total cytochrome P450 and reduced glutathione in rat hepatocytes/MS co-cultures were 31% and 127% respectively of the day O values, whereas they were undetectable in pure culture. A clear induction of monooxygenase activities by methylcholanthrene, phenobarbital and ethanol could be observed by the 5th day in MS cells/hepatocyte co-cultures. The fact that the results of our work show the suitability of MS cells, an epithelial-derived cell line, for improving the expression of biotransformation enzymes of cultured hepatocytes opens new possibilities of simplifying co-cultures for their use in drug-metabolism studies.

Possible prognostic value of pulmonary ah‐locus‐linked enzymes in patients with tobacco‐related lung cancer

As prognosis in breast cancer patients has been related to the AHH activity in their breast tissue, we have conducted a similar analysis on pulmonary drug metabolizing enzymes as prognostic markers for male lung cancer patients, primarily investigated for other reasons. A subset of 50 patients with lung cancer related to tobacco use, who had undergone thoracic surgery, was re-analyzed. The activity of parenchymal aryl hydrocarbon hydroxylase (AHH) and epoxide hydrolase (EH) that had been determined previously in homogenates of non-neoplastic surgical lung specimens, was used for comparisons of the patients' survival after surgery. When the crude mortality percentages at 1 and 2 years by AHH or EH activity, subdivided into quarters of the distribution, were calculated, a lower mortality was related to lower enzyme levels. Subjects in the 1st and 4th quarters of the distribution showed significant differences in their 1-year survival for AHH (p = 0.05) and EH (p less than 0.01) activities. This relationship could not be accounted for by age, cumulative lifetime smoking, recent or continuing smoking, stage or histological type of disease. Thus, the levels of pulmonary AHH and EH may have some prognostic significance in tobacco-related lung cancer.

Influence of the Ah locus on the humoral immunotoxicity of 2,3,7,8-tetrachlorodibenzo- p-dioxin: Evidence for Ah-receptor-dependent and Ah-receptor-independent mechanisms of immunosuppression

There are conflicting reports in the literature regarding the role of the Ah locus in 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) immunotoxicity. The present studies have utilized two congenic strains of C57Bl 6 mice that differ only at this locus to assess its influence on TCDD-induced suppression of antibody responses. Mice were given a single oral dose of TCDD 2 days prior to challenge with sheep red blood cells (SRBC) or trinitrophenyl-lipopolysaccharide (TNP-LPS). The subsequent dose-dependent effects of TCDD on the amount of antibody produced by splenic plasma cells were measured using the hemolytic antibody isotope release assay. In addition, the relative importance of the Ah genotype of lymphoid versus nonlymphoid tissue was examined in adoptive transfer experiments. Aryl hydrocarbon hydroxylase (AHH) activity was significantly induced in Ah bb mice by a dose of 0.5 μg/kg TCDD and maximally induced by a dose of 2 μg/kg. Ah dd mice required 10-fold higher doses of TCDD to induce comparable levels of AHH. The degree of thymic involution and liver hypertrophy induced by TCDD was also influenced by the Ah genotype of the animals. Both Ah bb and Ah dd mice exhibited dose-dependent suppression of the anti-TNP response following TCDD exposure. The ID50 was 7.0 μg/kg in Ah bb mice and 30.8 μg/kg in Ah dd mice. Suppression of the antibody response to SRBC was also dependent on the Ah locus. The ID50 in Ah bb mice was 0.6 μg/kg TCDD. However, an apparent biphasic dose response for suppression of the anti-SRBC response in Ah dd mice suggested the involvement of an Ah-independent component of suppression as well. In adoptive transfer studies, lymphocytes were identified as an Ah-dependent component of the response. The Ah-independent component of the response was not identified, and could be either lymphoid or nonlymphoid in nature. The possibility that T helper cells represent the Ah-independent component is discussed.

Aryl hydrocarbon hydroxylase activity levels in the hepatic microsomal fraction from MC- or TCDD-treated mice: a comparison between aromatic hydrocarbon responsive and non-responsive strains

The effects of in vivo administration of polycyclic aromatic hydrocarbons on the levels of aryl hydrocarbon hydroxylase (AHH) activity in aromatic hydrocarbon (Ah) responsive and non-responsive strains of mice were studied using the hepatic microsomal fraction. Injection of 3-methylcholanthrene (MC; 42 mg kg body wt.) or 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD; 120 μg kg body wt.) into both strains produced marked enhancement of AHH activity except for MC treatment of Ah non-responsive strains. Addition of 7,8-benzoflavone (BNF) to the microsomal AHH assay mixture prepared from mice previously injected with vehicle (olive oil) alone caused an increase in activity when the mice were responsive, while BNF lowered the activity in non-responsive strains. With regard to MC-injected mice, BNF and 3-methyl-sulphonyl-4,5,3',4'-tetrachlorobiphenyl (3-MSF-TCB) decreased microsomal AHH activity in Ah-responsive mice, whereas these drugs enhanced the activity in Ah-non-responsive strains. 3-MSF-TCB also had inhibitory potency on AHH activity, but the mechanism of inhibition seems to be somewhat different from that of BNF. It may also suggest that cytochrome P-450 isozymes inhibited by BNF are different from those inhibited by 3-MSF-TCB.

Variation in Inducibility of Cytochrome P-450c and Aryl Hydrocarbon Hydroxylase in Rat Liver, Lung, Kidney, Pancreas and Nasopharynx

The effect of 3-methylcholanthrene (MC) treatment on the cytochrome P-450c content of various rat tissues was examined by measuring the level of immunodetectable P-450c in conjunction with its aryl hydrocarbon hydroxylase (AHH) activity. Immunoblots revealed that P-450d was induced in the nasopharynx and pancreas in addition to its previously reported induction in the liver, lung and kidney. In contrast to P-450c, induction of the immunorelated P-450d was observed only in the liver. The specific content of immunodetected P-450c in the tissue homogenates decreased in the order: liver, nasopharynx, pancreas, lung, kidney. The corresponding AHH activities decreased in the order: liver, kidney, lung, nasopharynx, pancreas. The ratio of AHH activity to P-450c content varied widely among tissues: ratios of 37.2:1.7:0.47:0.04:0.02 were obtained for the kidney, liver, lung, nasopharynx and pancreas, respectively. The absence of a direct relationship between the levels of AHH and P-450c indicates that the extrahepatic activity may partially derive from P-450 forms other than P-450c and/or the specific activity of P-450c varies among different tissues.

Aryl hydrocarbon hydroxylase activity in human placenta of passive smokers

The levels of smoke components and metabolites in maternal blood and urine are useful in assessing direct exposure but they do not appear to be sufficiently sensitive as a long-term indicator of passive smoke exposure. Induction of aryl hydrocarbon hydroxylase (AHH) activity in the placenta as a result of maternal smoking has been well documented. This enzyme oxidizes various polycyclic aromatic hydrocarbons abundantly present in cigarette smoke. We hypothesized that passively inhaled tobacco smoke may induce placental AHH activity. Placental AHH levels were determined in 207 pregnancies at birth. As has been found in previous studies, we demonstrated that smoking during pregnancy is associated with a marked increase in placental AHH activity. A relationship was found between the recorded number of cigarettes smoked per day and the placental AHH activity. Moreover, AHH activity was significantly higher in pregnant women passively exposed to tobacco smoke relative to controls. The usefulness of analysis of placental AHH activity as a biological marker of in utero smoke exposure in epidemiological studies is considered.

2,3,7,8-Tetrachlorodibenzo- p-dioxin-induced porphyria in genetically inbred mice: Partial antagonism and mechanistic studies

2,3,7,8-Tetrachlorodibenzo- p-dioxin (TCDD) (233 nmol/kg) causes a significant increase of hepatic uroporphyrin, heptacarboxyporphyrin, and total porphyrins in female C57BL/6 mice, ovariectomized C57BL/6 mice, male C57BL/10 mice, and male C57BL/6 mice 3 weeks after treatment. In contrast, 6-methyl-1,3,8-trichlorodibenzofuran (MCDF) was inactive at a dose of 750 μmol/kg. Cotreatment of the mice with TCDD (233 mol/kg) plus MCDF (750 μmol/kg) resulted in partial antagonism of TCDD-induced hepatic porphyrin accumulation only in the female mice. Parallel studies in female C57BL/6 mice showed that the TCDD-induced porphyria was accompanied by the induction of hepatic microsomal aryl hydrocarbon hydroxylase (AHH) and ethoxyresorufin O-deethylase (EROD) activities and the depression of uroporphyrinogen decarboxylase (UROD). MCDF (750 μmol/kg) did not significantly affect these enzymes. In the cotreatment studies (MCDF plus TCDD), MCDF partially antagonized TCDD-induced hepatic porphyrin accumulation but did not affect the levels of hepatic AHH, EROD, or UROD. These results indicate that other factors, in addition to the induction of cytochrome P450-dependent monooxygenases and depressed UROD activity, are important in TCDD-induced porphyria in C57BL/6 female mice.

Drug-metabolizing enzymes in rat, mouse, pig and human macrophages and the effect of phagocytic activation

Aryl hydrocarbon hydroxylase (AHH) activity mediated by cytochrome P-450 is present in pig hepatic microsomes [10 nmol · 3 mg protein −1 · hr −1]. AHH activity was detectable in both hepatocytes and Kupffer cells isolated from pig liver biopsy material. These cells were isolated from needle or wedge biopsy material by collagenase perfusion and incubation with collagenase at 37°. The two cell types were separated from the resulting cell suspension as previously described for whole liver. Kupffer cells were enriched by adherence and were cultured for 24 hr prior to harvesting. Cells were harvested, and cell viability was determined. AHH activity was assayed in Kupffer cell and hepatocyte homogenates. Kupffer cell AHH activity was approximately one-eighth the level detected in hepatocytes. To determine whether this enzyme was present in other macrophages, monocytes were isolated from 10 ml of heparinized peripheral blood using Ficoll-Hypaque and were enriched by adherence. After 24 hr in culture, cell viability was assessed and monocytes were identified by cytochemical staining. AHH activity was detectable in pig monocyte homogenates, and the AHH level was similar to that in pig Kupffer cells. AHH was also easily detectable in human monocytes. This macrophage AHH activity was compared with AHH activity in rat monocytes, mouse Kupffer cells and mouse peritoneal macrophages. Monocyte AHH was relatively stable in cell culture but decreased rapidly upon storage at −70°. Macrophage AHH activity was depressed following phagocytic activation in vitro by latex beads with a concomitant increase in heme oxygenase activity.

Effectiveness of a Prudhoe Bay crude oil and its aliphatic, aromatic and heterocyclic fractions in inducing mortality and aryl hydrocarbon hydroxylase in chick embryo in ovo.

Prudhoe Bay crude oil (PBCO) and its aliphatic, aromatic and heterocyclic fractions were tested on the developing chick embryo for (i) embryotoxicity (ii) their ability to induce hepatic and renal cytochrome P450 levels as well as hepatic, renal and pulmonary aryl hydrocarbon hydroxylase activities. On the basis of its concentration in PBCO, the aromatic fraction was responsible for most of the embryotoxicity as well as for the enzyme inducing ability. The NOS fraction constituted less than 7% (w/v) of PBCO but, on a weight equivalent basis, was roughly as potent as the aromatic fraction in causing embryotoxicity and in inducing cytochrome P450 levels and aryl hydrocarbon hydroxylase. The aliphatic fraction was found to be essentially inactive. The results are consistent with the concept that elevation of aryl hydrocarbon hydroxylase levels by certain components of PBCO may lead to increased embryotoxicity.

Induced Cytochrome P450 in Winter Flounder (Pseudopleuronectes americanus) from Coastal Massachusetts Evaluated by Catalytic Assay and Monoclonal Antibody Probes

Levels of hepatic microsomal ethoxyresorufin O-deethylase (EROD) and aryl hydrocarbon hydroxylase (AHH) activities and sensitivity to inhibition by α-naphthoflavone in winter flounder (Pseudopleuronectes americanus) from Deer Island flats in Boston Harbor, off Plymouth Beach, off Nantucket, and at the outer New Bedford Harbor in Buzzards Bay, Massachusetts, suggested induction of cytochrome P450 by environmental chemicals. Levels of activity were higher in fish from Boston Harbor and Plymouth Bay than from Nantucket and Buzzards Bay. In fish from Buzzards Bay the levels of EROD and AHH activities were closely correlated, with some fish there apparently uninduced. Immunoblot analysis of flounder liver microsomes with a monoclonal antibody (1-12-3) against β-naphthofiavone-inducible scup cytochrome P450E revealed a single cross-reacting protein in untreated fish from all four field sites. A similar protein was induced by β-naphthoflavone treatment. Levels of this flounder protein correlated positively with levels of AHH and EROD activity in Buzzards Bay fish, consistent with a conclusion that some fish there were uninduced. The results demonstrate the induction of a P450E counterpart in flounder in Massachusetts waters and indicate its identity as the AHH and EROD catalyst. The results also establish the use of monoclonal anti-P450E in analysis of environmental induction.