Cytochrome P4501A Response Research Articles

Interindividual variation in the cytochrome P4501A response to 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin in herring gull embryo hepatocytes

Exposure to dioxin-like compounds is consistently associated with concentration-dependent induction of cytochrome P4501A (CYP1A) enzymes in primary cultures of avian hepatocytes. We have previously demonstrated that the median effective concentration (EC50) for induction of this response is predictive of in vivo sensitivity to dioxin-like compounds in birds. We investigated sources of interindividual variation in the CYP1A response to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in wild herring gulls and considered how this variation may complicate dioxin sensitivity estimates based on the CYP1A bioassay. Concentration-dependent effects of TCDD on CYP1A mRNA expression were characterized in 55 hepatocyte cultures prepared from individual herring gull embryos. A large degree of variability was observed among the hepatocyte culture preparations. For example, 1) basal CYP1A4 and CYP1A5 mRNA expression varied by 20- and 126-fold, respectively, among individuals, and 2) exposure to TCDD induced CYP1A4 mRNA expression by 57-fold in the most responsive sample but did not significantly induce CYP1A4 mRNA expression above baseline values in 42% of hepatocyte culture preparations. Environmental and genetic factors contributing to the observed variability are discussed. Despite the large amount of interindividual variation, we conclude that reproducible EC50-based estimates of species sensitivity can be obtained from the CYP1A cell culture bioassay when samples are collected from relatively uncontaminated colonies. Environ Toxicol Chem 2019;38:660-670. © 2019 SETAC.

Expression of aryl hydrocarbon receptor-regulated genes and superoxide dismutase in the Antarctic eelpout Pachycara brachycephalum exposed to benzo[a]pyrene.

The aryl hydrocarbon receptor (AhR) pathway mediates many, if not all, responses of fish to dioxin-like compounds. The Southern Ocean is progressively exposed to increasing concentrations of anthropogenic pollutants. Antarctic fish are known to accumulate those pollutants, yet nothing is known about their capability to induce chemical biotransformation via the AhR pathway. The objective of the present study was to investigate whether Antarctic eelpout, Pachycara brachycephalum, respond to anthropogenic pollutants by activation of the AhR and its target gene cytochrome P4501A (CYP1A), and of superoxide dismutase (SOD), which served as a representative for oxidative stress. We exposed P. brachycephalum to 10 and 100 mg benzo[a]pyrene (BaP)/kg body weight for 10 d and measured the expression of AhR, CYP1A, and SOD in liver tissue via quantitative polymerase chain reaction. We identified two distinct AhR isoforms in the liver of P. brachycephalum. Antarctic eelpout responded to both BaP exposures by an up-regulation of AhR and SOD, and by a particularly strong induction of CYP1A expression, which remained high until day 10 of the exposure time. Our data suggest that P. brachycephalum possesses the potential to up-regulate xenobiotic biotransformation pathways, at least at the gene expression level. The time course of the AhR and CYP1A response points to an efficient but slow xenobiotics metabolism. Moreover, BaP exposure could include adverse effects such as oxidative stress. Environ Toxicol Chem 2018;37:1487-1495. © 2018 SETAC.

CYP1A expression in liver and gills of roach (Rutilus rutilus) after waterborne exposure to two phenanthrene derivatives, 1‐methylphenanthrene and 4‐methylphenanthrene

Phenanthrenes (Phs) substituted with alkyl groups are a class of compound present in the environment, and they appear to be toxic to developing fish. The present study aimed to investigate the effect of waterborne exposure to two monomethyl derivatives of phenanthrene, 1-methylphenanthrene (1M-Ph) and 4-methylphenanthrene (4M-Ph), on cytochrome P450 1A (CYP1A) gene expression in fish gills and liver. Juvenile common roaches (Rutilus rutilus) were exposed to water with dimethyl sulfoxide (DMSO) solutions of 1M-Ph, 4M-Ph, benzo[a]pyrene (BaP; positive control), each at a dose of 100 µg/L, or to water with DMSO alone (negative control group) for 2 d and 7 d. Significant CYP1A responses with regard to treatment and exposure duration were noted (2-way analysis of variance [ANOVA]) in gills (p = 0.013 and p = 0.003, respectively) and liver (p < 0.001). The 2 monomethyl Phs did not induce consistent gene expression changes, except for 4-MPh, which elevated the CYP1A messenger ribonucleic acid (mRNA) level in the liver at the end of the treatment (almost 4-fold; p < 0.05; 7 d). As was expected, exposure to BaP resulted in elevation of CYP1A mRNA expression in treated fish compared with the control group. Expressions after 2 d and 7 d were approximately 220- and 180-fold higher in liver and 8- and 6-fold higher in gills respectively. The CYP1A protein levels remained stable in both tissues, with one notable exception in roach liver treated for 2 d with BaP (∼ 6-fold increase; p < 0.05). The different effects of the 1- and 4-methylphenanthrenes on CYP1A gene expression in roach liver suggest a relationship between chemical or 3-D structure of the differentially substituted monomethyl Phs and their biological activity.

Lack of CYP1A responsiveness in species inhabiting chronically contaminated habitats: Two varieties of resistance?

Organisms chronically exposed to organic pollutants such as polychlorinated biphenyls (PCBs) can develop resistance to these chemicals, a condition associated with reduced inducibility of the biomarker enzyme cytochrome P450 1A (CYP1A). This study addresses the CYP1A response of members of the families Ictaluridae and Centrarchidae, two fish families found throughout much of the United States. We measured CYP1A expression, PCB body burdens, and conducted CYP1A challenge experiments in species from these families residing in the Town Branch/Mud River system (Logan County, KY, USA), a stream system historically contaminated with high levels of PCBs. Despite PCB concentrations in muscle tissue typically associated with elevated CYP1A (16.7 to 75.2μgPCB/g wet edible flesh), resident fish in the contaminated Town Branch/Mud River sites (yellow bullhead [Ameiurus natalis], green sunfish [Lepomis cyanellus], and spotted bass [Micropterus punctulatus]) had hepatic CYP1A activity levels similar to, rather than higher than, those in reference fish, suggesting reduced sensitivity to CYP1A induction. Lack of CYP1A expression following direct contaminant exposure has often been associated with resistance to those contaminants. To determine if CYP1A in resident populations was resistant to induction by PCBs, we exposed resident fish to a single, intraperitoneal injection with a potent CYP1A inducer, 3,4,3′,4′-tetrachlorobiphenyl (PCB 77). PCB 77 treatment significantly induced hepatic CYP1A activity and protein in yellow bullhead from reference, but not contaminated, sites and had no effect on CYP1A in green sunfish from either site. The low CYP1A expression levels in resident fish with elevated PCB body burdens, together with the failure of PCB injection to induce CYP1A in certain populations, indicate an acclimatory CYP1A response in yellow bullheads and likely an inherently resistant CYP1A in green sunfish. This work demonstrates for the first time acclimation of CYP1A to PCBs in a species within the family Ictaluridae and provides further support for our previous work indicating an apparent inherent lack of CYP1A sensitivity to chlorinated inducers in Centrarchids. These traits may explain, at least in part, the common association of these families with degraded habitats and indicate Lepomis members are likely to be excellent candidates for exploring the mechanistic basis of ‘inherent’ CYP1A resistance. This study also underlines to the need for thorough characterization of the CYP1A responsivity of a population and/or species prior to using CYP1A as a reliable biomonitoring tool.

Differential sensitivity of CYP1A to 3,3′,4′,4-tetrachlorobiphenyl and benzo(a)pyrene in two Lepomis species

Although Lepomis species are abundant in a wide variety of habitats throughout North America and could serve as potentially valuable biomonitoring tools, few studies have examined the induction of pollutant biomarkers in this genus. We hypothesized that the induction of cytochrome P-450 1A (CYP1A), a sensitive and widely used indicator of response to aquatic contaminants, would serve as an effective biomarker of organic pollutant exposure in Lepomis species. We examined the response of CYP1A and two of the major pollutant-responsive phase II enzymes, glutathione S-transferase (GST), and uridine diphosphate glucuronyltransferase (UDPGT), in Lepomis exposed to organic pollutants under laboratory and field conditions. Two Lepomis species (longear sunfish, Lepomis megalottis and bluegill, Lepomis macrochirus) were exposed in the laboratory via intraperitoneal injection to corn oil (vehicle), benzo(a)pyrene (BaP) (10 and 50 mg/kg), a polynuclear aromatic hydrocarbon (PAH) or 3,4,3′,4′-tetrachlorobiphenyl (PCB 77) (0.1 and 1.0 mg/kg), a dioxin-like planar halogenated aromatic hydrocarbon (HAH), and sacrificed 2 (BaP) or 7 (corn oil, PCB77) days later. Lepomis hepatic CYP1A exhibited differential sensitivity to these two classes of environmental contaminants. CYP1A activity was weakly induced in bluegill exposed to 1.0 mg/kg PCB 77 (3 fold induction over controls) but strongly induced in both bluegill and longear sunfish exposed to 50 mg/kg BaP (37 and 15 fold induction over controls, respectively). In contrast, hepatic GST activity in both species remained unchanged following the treatment with either compound and hepatic UDPGT activity, which was assessed only in BaP-treated longear sunfish, was unaffected by that chemical, indicating these phase II enzymes may not be sensitive pollutant biomarkers in this genus. Further, longear sunfish collected from a PCB contaminated site displayed relatively low levels of CYP1A activity despite PCB body burdens associated with strong induction of CYP1A activity in other fish species. The strong induction of CYP1A by BaP with much weaker CYP1A response to PCB indicates that CYP1A in Lepomis sp. could be an excellent biomarker for PAH pollution, but may not be a reliable indicator of site contamination by halogenated hydrocarbons. We conclude that Lepomis species provide a useful model for examining the regulation and potential consequences of differential pollutant sensitivity, but that CYP1A in these species should be used with caution as an indicator of halogenated contaminants.

CYP1A Expression in Caged Rainbow Trout Discriminates Among Sites with Various Degrees of Polychlorinated Biphenyl Contamination

It has become increasingly apparent that resident fish can develop resistance to chemicals in their environment, thus compromising their usefulness as sentinels of site-specific pollution. By using a stream system whose resident fish appear to have developed pollutant resistance (Brammell et al., Mar Environ Res 58:251-255, 2005), we tested the hypothesis that the pollutant-inducible biomarker, cytochrome P4501A (CYP1A), as measured in field-caged juvenile rainbow trout (Oncorhynchus mykiss), would reflect relative pollution differences between reference and polychlorinated biphenyl (PCB)-contaminated sites. Trout were caged in the Town Branch/Mud River system (Logan County, KY), a stream system undergoing remediation for PCBs. Fish were held in remediated (Town Branch), unremeditated (Mud River), and reference sites for 2 weeks during spring 2002. At the end of this period, gill and hepatic CYP1A expression were measured. To evaluate the relative PCB exposure of caged trout and provide a reference point against which to calibrate CYP1A response, PCB levels were quantified in sediments from each site. Hepatic CYP1A expression in caged trout clearly detected the presence of PCBs in the Town Branch/Mud River stream system. Sediment PCB levels and hepatic CYP1A expression in caged trout produced identical pollution rankings for the study sites. Gill CYP1A expression, although suggestive of site differences, was not statistically different among sites. Unlike resident fish, which failed to show site differences in hepatic CYP1A expression in this waterway (Brammell et al. 2005), caged fish proved to be a sensitive discriminator of relative PCB contamination in this system. In summary, we determined that CYP1A expression in caged fish reflected relative in situ pollutant exposure. The exposure paradigm confirmed that 2 weeks was a sufficient caging period for evaluating CYP1A response in this species at these temperatures (13-19 degrees C). In addition, these studies demonstrate that tissue-specific CYP1A expression can provide insights into likely routes of exposure. We conclude that CYP1A expression in caged trout is a reliable and inexpensive first-pass determination of relative environmental pollutant exposure and bioavailability in aqueous systems.

Influence of 17α-ethynylestradiol on CYP1A, GST and biliary FACs responses in male African sharptooth catfish (Clarias gariepinus) exposed to waterborne Benzo[a]Pyrene

Dose dependent effects of Benzo[a]pyrene (BaP) on cytochrome P4501A (CYP1A), glutathione-S-transferase (GST) and fluorescent aromatic compounds (FACs) metabolites biomarker responses were studied in African sharptooth catfish (Clarias gariepinus) following 24 h of waterborne exposures. Based on biomass of C. gariepinus in different tanks, BaP concentrations of 1.60, 3.44, and 18.21 microg/L that corresponded to 0.5, 1.0, and 5.0 mg/kg body weight were used. Significant induction of EROD activities in gill filaments was observed at all doses and the accumulation of FACs metabolites in bile was significantly different between groups. Accumulation of FACs metabolites in bile strongly correlated (r (2) = 0.99) with BaP doses. Hepatic EROD activities were undetectable and no effect on GST activities was observed. The highest dose of BaP from the dose dependent study was further studied to assess the interactive and temporal responses of C. gariepinus on CYP1A, GST, and FACs metabolites biomarkers following exposure to either BaP alone, 17alpha-ethynylestradiol (EE(2)) alone or a combination of both compounds at concentrations of 54.17 microg/L for BaP, 51.38 microg/L for EE(2) and 54.44 microg/L for each of both compounds. Based on biomass in each tank, these concentrations corresponded to 5 mg/kg body weight. While a group of six fish was sacrificed on day 0 from the control tank only, other groups of six fish were sacrificed after 1, 3, and 6 days of exposure from the control and exposed groups. Maximum induction of gill filament and hepatic EROD activities was observed after 1 day of exposure. Both EROD activities in gill filaments and liver were significantly induced by exposure to BaP alone or co-administration with EE(2). Gill filament EROD induction was significantly inhibited (50%) by co-administration of BaP and EE(2) compared to administration of BaP alone. Levels of FACs in bile for BaP and BaP + EE(2) exposed groups were significantly different from the control at all doses and time points. A significant induction of GST activities was observed in fish exposed to BaP and BaP + EE(2) after 3 days. Exposure to EE(2) alone caused significant induction of this enzyme after day 6. This study reports for the first time the significant antagonistic influence of EE(2) on BaP in gills of fish following waterborne exposures. The results also indicate that chemical mixtures may affect biomarker responses differently from compounds administered alone and that the sensitivity of CYP1A to interactive chemicals is different in gills and liver.

Exposure to 3,3′,4,4′,5‐pentachlorobiphenyl during embryonic development has a minimal effect on the cytochrome P4501A response to 2,3,7,8‐tetrachlorodibenzo‐P‐dioxin in cultured chicken embryo hepatocytes

Concentrations of dioxin-like compounds in avian eggs can vary substantially between individuals, species, and collection sites. Although the inducibility of cytochrome P4501A (CYPIA) in hepatocyte cultures is commonly used to predict species sensitivity to environmental contaminants, it is not known how exposure to dioxin-like compounds during embryonic development might alter this biomarker response. To investigate this question, we injected vehicle or 0.4, 0.8, or 1.6 microg/kg of 3,3',4,4',5-pentachlorobiphenyl (PCB 126) into the air cell of fertilized chicken eggs before incubation, and we measured CYP1A endpoints in cultured hepatocytes of day-19 embryos from each treatment group. The CYP1A response to PCB 126 also was assessed in whole liver tissue. Embryonic exposure to the most environmentally relevant treatment, 0.4 microg/kg of PCB 126, increased CYPIA4 mRNA expression 29-fold compared to control values in whole liver tissue but only twofold compared to control values in cultured hepatocytes. The CYPIA response to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) was not altered in hepatocytes cultured from embryos treated with 0.4 microg/kg of PCB 126, but at 0.8 and 1.6 microg/kg of PCB 126, several concentration-dependent effects were observed. For example, embryos exposed to higher concentrations of PCB 126 in ovo were more responsive to CYP1A mRNA induction by TCDD in vitro. These findings suggest that exposure to environmental levels of dioxin-like compounds during incubation is not likely to alter species-sensitivity estimates derived from in vitro CYP1A data.

Fasting Augments PCB Impact on Liver Metabolism in Anadromous Arctic Char

Anadromous arctic char (Salvelinus alpinus) undertake short feeding migrations to seawater every summer and accumulate lipids, while the rest of the year is spent in fresh water where the accumulated lipid reserves are mobilized. We tested the hypothesis that winter fasting and the associated polychlorinated biphenyls' (PCBs) redistribution from lipid depots to critical tissues impair the liver metabolic capacity in these animals. Char were administered Aroclor 1254 (0, 1, 10, and 100 mg/kg body mass) orally and maintained for 4 months without feeding to mimic seasonal winter fasting, while fed groups (0 and 100 mg Aroclor 1254/kg) were maintained for comparison. A clear dose-related increase in PCB accumulation and cytochrome P4501A (CYP1A) protein content was observed in the livers of fasted fish. This PCB concentration and CYP1A response with the high dose of Aroclor were 1.5-fold and 3-fold greater in the fasted than in the fed fish, respectively. In fed fish, PCB exposure lowered liver glycogen content, whereas none of the other metabolic indicators were significantly affected. In fasted fish, PCB exposure depressed liver glycogen content and activities of glucose-6-phosphate dehydrogenase, alanine aminotransferase, lactate dehydrogenase, and phosphoenolpyruvate carboxykinase and elevated 3-hydroxyacylcoA dehydrogenase activity and glucocorticoid receptor protein expression. There were no significant impacts of PCB on heat shock protein 70 (hsp70) and hsp90 contents in either fed or fasted fish. Collectively, our study demonstrates that winter emaciation associated with the anadromous lifestyle predisposes arctic char to PCB impact on hepatic metabolism including disruption of the adaptive metabolic responses to extended fasting.

Cellular distribution and induction of CYP1A following exposure of gilthead seabream, Sparus aurata, to waterborne and dietary benzo(a)pyrene and 2,3,7,8-tetrachlorodibenzo- p-dioxin: An immunohistochemical approach

The present study aimed to investigate cellular expression of cytochrome P4501A (CYP1A) protein in the seabream, Sparus aurata, exposed to one of two CYP1A-inducing contaminants, benzo(a)pyrene (B(a)P) or 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD). Male adult fish were exposed to several concentrations of TCDD or B(a)P either via water or via food. Fish were sampled after 0, 5, 10, 15 or 20 days of treatment and the time- and concentration-dependent induction of CYP1A protein in cells and tissues was studied using immunohistochemistry. A general site of CYP1A induction was the vascular endothelium. Aqueous exposures resulted in elevation of CYP1A immunoreactivity in gill pillar cells, heart endothelium, renal tubular epithelium, hepatocytes, and gut mucosal epithelium. In contrast, dietary exposure resulted in strong CYP1A immunostaining in gut epithelium but in only mild to moderate staining elsewhere. Both B(a)P and TCDD induced CYP1A in similar cellular response patterns in most organs examined, although TCDD generally led to higher staining intensity and frequency (i.e. the number of CYP1A-positive cells within an organ), an effect that is likely to be related to compound-specific differences in induction potency, metabolism and penetration. Contaminant-specific staining patterns were observed in the gills, where TCDD exposure evoked CYP1A immunostaining in the endothelial pillar cells, while B(a)P induced CYP1A staining in the branchial epithelial cells. This work points to the importance of immunohistochemical identification of cell-specific CYP1A responses in assessing the toxicology of CYP1A-inducing xenobiotics.

Cytochrome P4501A induction in brown trout exposed to small streams of an urbanised area: results of a five-year-study

This case study examines the ability of the cytochrome P4501A (CYP1A) biomarker to distinguish the pollution status of two small streams, Krähenbach and Körsch, receiving different levels of urban and agricultural impact, with low to moderate contamination by arylhydrocarbon receptor (AhR)-binding PAHs and PCBs. Brown trout, Salmo trutta, exposed in enclosure restrictions, showed significant between-stream differences of hepatic CYP1A levels. EROD activities were the better discriminator than CYP1A protein levels. The CYP1A response was consistent and repeatable over the 5-year observation period from 1995 to 1999. In contrast to brown trout, hepatic CYP1A of stone loach, Barbatula barbatula, did not clearly distinguish the streams. The findings of this long-term study lend support to the use of CYP1A as a biomarker of degraded environmental conditions, provided that sufficiently long observation periods are used to average out confounding factors, that sufficiently sensitive detection methods are used, and that a responsive monitoring species is chosen.

Cytochrome P4501A (CYP1A) in killifish (Fundulus heteroclitus): heritability of altered expression and relationship to survival in contaminated sediments.

Previous research has shown that killifish (Fundulus heteroclitus) inhabiting a creosote-contaminated site on the Elizabeth River in Virginia exhibit little induction of cytochrome P4501A (CYP1A) protein expression and activity upon exposure to typical CYP1A-inducing chemicals. We characterized the CYP1A response of first, second, and third generation laboratory-raised offspring of feral Elizabeth River killifish to exposure to sediments from the contaminated site as well as the prototypical polycyclic aromatic hydrocarbon (PAH)-type CYP1A inducers beta-naphthoflavone (BNF) and 3-methylcholanthrene (3-MC). The Elizabeth River offspring's responses were compared to those of offspring of killifish from two reference sites (King's Creek, Virginia, and Russell Creek, North Carolina). As with feral Elizabeth River killifish, the first generation embryos and larvae were refractory to CYP1A induction. However, the response observed in 3-year-old first generation adults, as well as with second and third generation fish, was much closer to that observed in reference-site fish. We suggest that the pattern of altered CYP1A response in Elizabeth River killifish, while persistent and heritable for one generation, is mostly nongenetically based. Additionally, we investigated the hypothesis that low CYP1A activity (measured as in ovo EROD activity) would correlate to increased survival in Elizabeth River sediment pore water; this hypothesis was not supported by our results.

Cytochrome P4501A responses in two tropical fish species, riverine catfish ( Rita rita) and marine mudfish ( Apocryptes bato)

The present study focuses on the establishment of methods for biomarker studies in freshwater and marine fish species as a basis for monitoring the extent of contamination of fisheries resources in tropical waters. Riverine catfish ( Rita rita) and marine mudfish ( Apocryptes bato) were given a single intraperitoneal injection with two selected inducing compounds; β-naphthoflavone (BNF, 50 mg/kg) and a polychlorinated biphenyl mixture (Clophen A50, 20 mg/kg), and the heavy metal compound cadmium chloride (CdCl 2, 1 mg/kg). Effects on cytochrome P4501A (CYP1A) were determined in post-mitochondrial supernatants (PMS) of liver at days 3 and 10 after treatment. EROD (7-ethoxyresorufin O-deethylase) activity and CYP1A protein level by indirect non-competitive enzyme-linked immunosorbent assay (ELISA) and Western blotting using a monoclonal antibody against fish CYP1A, were measured. BNF and Clophen A50 resulted in induction upto 9.5- and 5-fold, respectively, of CYP1A protein compared to control, while CdCl 2 showed significant inhibition in these species. The present study examined the phase-I cytochrome P450 monooxygenase activity and response in these two tropical fish species for the first time.

Confinement stress and starvation modulate the induction of EROD activity after dietary exposure to PCB 126 in the Mozambique tilapia (Oreochromis mossambicus)

Polychlorinated biphenyls (PCBs) are environmental pollutants, accumulating in the food chain and inducing cytochrome P450 1A (CYP P450 1A) monooxygenase enzymes. This study aimed to investigate the influence of dietary PCB 126 exposure on cytochrome P4501A-associated enzyme activities in head kidney, liver, gill and intestine of unstressed and stressed tilapia (Oreochromis mossambicus) before and after starvation. Fish were fed diets containing 0 μg (control), 0.5 μg (low) or 50 μg (high) PCB126 per kg body weight per day for 7 days. After the PCB-exposure each treatment group was sampled directly, or was subsequently exposed to confinement stress. Replicate groups remained undisturbed, starved for 3 weeks and were thereafter sampled directly or after confinement. Catalytic activity of CYP P4501A was determined as 7-ethoxyresorufin-O-deethylase (EROD) activity. Directly after PCB exposure and after starvation, EROD activities in liver, gill and head kidney increased with higher PCB dose. Intestinal EROD activities were not detectable at any sampling point. Confinement, known to evoke a stress response increased EROD activities in livers (only directly after PCB exposure) and in head kidneys (directly after PCB exposure and after starvation) but only in fish fed the low PCB-diet. We suggest that stressing tilapia induces EROD activities at non-saturating PCB concentrations and that this effect is more pronounced in nourished than in starved fish. These data show that the cytochrome P450 1A response to toxicants in fish is also influenced by non-chemical factors. This should be considered when EROD activities are used as a biomarker for environmental monitoring studies.

Estrogenic and CYP1A response of mummichogs and sunshine bass to sewage effluent

Recent studies demonstrating feminization of effluent-exposed wild-caught male fish in the UK have prompted much research regarding the estrogenic activity of effluent from municipal sewage treatment plants (MSTPs). To investigate the estrogenicity and cytochrome P450 1A (CYP1A) induction potency of MSTP effluent, two species of fish, adult male mummichogs, Fundulus heteroclitus, and juvenile sunshine bass, Morone saxatilis × Morone chrysops, were exposed to un-chlorinated effluent (75% effluent, 25% seawater) from a large MSTP in Yonkers, NY, USA. After a 21-day static-daily (75%) renewal exposure, significant elevations over controls were observed in levels of vitellogenin (VtG) in plasma (1730%) and liver (131%) in effluent-exposed sunshine bass. In contrast, hepatic VtG was not elevated in mummichogs; plasma VtG was not measured in this species. Effluent exposure elevated hepatic CYP1A protein (140–145%) and ethoxyresorufin- O-deethylase (EROD) activity (408–598%) in both species. These findings suggest ontogenetic and/or species differences in response to estrogenic compounds in MSTP effluent. Furthermore, the elevation of CYP1A in response to sewage effluent exposure indicates the presence of additional compounds that may alter xenobiotic and/or steroid biotransformation in fish.

Environmental contamination and developmental abnormalities in eggs and hatchlings of the common snapping turtle ( Chelydra serpentina serpentina) from the Great Lakes—St Lawrence River basin (1989–1991)

During 1989–1991, we assessed developmental abnormalities in embryos and hatchlings from eggs of the common snapping turtle ( Chelydra serpentina serpentina). Eggs were collected and artificially incubated from eight sites in Ontario, Canada and Akwesasne/New York, USA. In eggs from the same clutches we measured 20 organochlorine pesticides, 48 polychlorinated biphenyl (PCBs) congeners including 6 non-ortho PCBs, 8 polychlorinated dibenzodioxins (PCDDs), 14 polychlorinated dibenzofurans (PCDFs) and total mercury. We found a significant increase in abnormal development with increasing polychlorinated aromatic hydrocarbon exposure in eggs, particularly PCDD and PCDF concentrations. In contrast, the risk of abnormality was not significantly higher as toxic equivalent concentrations increased in eggs. We also found significant 7-ethoxyresorufin O-deethylase and Cytochrome P4501A responses in livers of hatchling turtles from Lake Ontario relative to hatchlings from a clean, inland site whereas we did not find any evidence of porphyria in the hatchlings from either site.

Cytochrome P450 1A response in North Sea Dab, Limanda limanda, from offshore and coastal sites

Dab ( Limanda limanda) were collected at four stations in the southern North Sea in November 1993. Highest muscle polychlorinated biphenyl (PCB) concentrations were found at a station near the Dutch coast. CB153 was the dominant congener at all stations. Since the concentrations of two mono- ortho Cl-substituted congeners covaried strongly with CB153, this congener appears to be a good marker for general differences in PCB concentrations. Mean CB153 concentrations ranged from 50–160 ng g −1 lipid. The highest cytochrome P450 1A (CYP1A) levels were also found at the coastal station. The extent of induction, measured catalytically (7-ethoxyresorufin O-deethylase, EROD) and immunochemically (CYP1A ELISA), between the less contaminated offshore sites and the coastal site ranged up to five-fold. Both parameters were well correlated with muscle tissue PCB concentrations. The data indicate that the CYP1A induction response in dab populations along the Dutch coast is strong enough to separate the Dutch coastal area from more pristine offshore areas of the North Sea. Compared to results of previous studies in other seasons the present sampling period offered the best conditions to investigate the correlation between environmental contamination with PCBs and related compounds, since the variations in natural factors that also interfere, such as water temperature and condition of the fish, were very low.

Use of cytochrome P450 1A (CYP1A) in fish as a biomarker of aquatic pollution.

Development of diagnostic and predictive biomarkers for use in pollution monitoring depends on a fundamental characterization and understanding of the mechanisms and regulations of the molecular response. Since first proposed as a pollution indicator in the mid-1970s (Payne & Penrose, 1975; Ahokas et al., 1975; Payne, 1976), the insight in the cytochrome P450 (CYP) system in fish, its molecular basis, regulation, and the mechanisms underlying the induction response to organic xenobiotics, has increased tremendously. Although the picture is still far from complete, we now have the knowledge and experience to put its potential as a pollution biomarker into use. However, understanding the influences of factors such as water temperature, season, sexual maturation, developmental status and diet, is an important and critical contribution to the full implementation of CYP1A responses in fish as a biomarker of aquatic pollution.

Use of cytochrome P450 1A (CYP1A) in fish as a biomarker of aquatic pollution

Development of diagnostic and predictive biomarkers for use in pollution monitoring depends on a fundamental characterization and understanding of the mechanisms and regulations of the molecular response. Since first proposed as a pollution indicator in the mid-1970s (Payne & Penrose, 1975; Ahokas et al., 1975; Payne, 1976), the insight in the cytochrome P450 (CYP) system in fish, its molecular basis, regulation, and the mechanisms underlying the induction response to organic xenobiotics, has increased tremendously. Although the picture is still far from complete, we now have the knowledge and experience to put its potential as a pollution biomarker into use. However, understanding the influences of factors such as water temperature, season, sexual maturation, developmental status and diet, is an important and critical contribution to the full implementation of CYP1A responses in fish as a biomarker of aquatic pollution.