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Gill EROD Activity in Fish: A Biomarker for Waterborne Ah-receptor Agonists
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Physiology and Developmental Biology, Environmental Toxicology.
2007 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Induction of the cytochrome P450(CYP)1A protein and the connected increase in 7-ethoxyresorufin O-deethylase (EROD) activity are common biomarkers in fish. Enhanced activity of this protein signals exposure to Ah-receptor agonists such as chlorinated dioxins, co-planar polychlorinated biphenyls (PCBs) and certain polycyclic aromatic hydrocarbons (PAHs). The EROD biomarker is commonly analyzed in liver microsomes. However, the gill is directly exposed to waterborne pollutants, and in this thesis the gill filament EROD assay was therefore evaluated as a monitoring tool for waterborne CYP1A inducers in fish. Originally developed in rainbow trout (Oncorhynchus mykiss), the assay was here applied in various limnic and marine species. Following exposure to low waterborne concentrations of the readily metabolized CYP1A inducers benzo(a)pyrene (BaP) and indigo, a strong EROD induction was observed in the gill but not in the liver. This likely reflected metabolic clearance of the inducers in gill and other extrahepatic tissues. The high sensitivity of the gill was confirmed in studies of fish caged in waters in urban and rural areas in Sweden where the gill consistently showed a more pronounced EROD induction compared with the liver and the kidney. Fish caged in the reference waters showed surprisingly strong gill EROD induction and CYP1A immunostaining. Consequently, there may be CYP1A inducers present in the aquatic environment that are not yet identified. The assay was further applied in Atlantic cod (Gadus morhua) as a biomarker of exposure to crude oil and produced water (PW) from oil fields in the North Sea. The assay was finally adapted to detect inhibiting compounds, and an imidazole, a triazole and a plant flavonoid turned out to be potent gill EROD inhibitors. The overall conclusion from the studies of this thesis is that the gill filament EROD assay is a practical and sensitive biomarker of exposure to waterborne CYP1A inducers in various fish species. The induction of gill EROD activity in fish also at the reference sites in the field studies calls for further studies on background contamination in Swedish waters.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis , 2007. , p. 52
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 311
Keywords [en]
Biology, biomarker, fish, gill, CYP1A, EROD, environmental monitoring, induction, inhibition, PAH, PCB, produced water
Keywords [sv]
Biologi
Identifiers
URN: urn:nbn:se:uu:diva-7899ISBN: 978-91-554-6902-3 (print)OAI: oai:DiVA.org:uu-7899DiVA, id: diva2:170305
Public defence
2007-06-02, Lindahlsalen, Evolutionsbiologiskt centrum (EBC), Norbyvägen 18 A, Uppsala, 10:00 (English)
Opponent
Supervisors
Available from: 2007-05-09 Created: 2007-05-09 Last updated: 2009-04-02Bibliographically approved
List of papers
1. EROD activity in gill filaments from anadromous and marine fish as a biomarker of dioxin-like pollutants
Open this publication in new window or tab >>EROD activity in gill filaments from anadromous and marine fish as a biomarker of dioxin-like pollutants
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2003 (English)In: Comparative Biochemistry and Physiology Part C, Vol. 136, p. 235-243Article in journal (Refereed) Published
Identifiers
urn:nbn:se:uu:diva-95920 (URN)
Available from: 2007-05-09 Created: 2007-05-09 Last updated: 2009-04-02Bibliographically approved
2. Cytochrome P4501A induction in rainbow trout gills and liver following exposure to waterborne indigo, benzo(a)pyrene and 3,3',4,4',5-pentachlorobiphenyl
Open this publication in new window or tab >>Cytochrome P4501A induction in rainbow trout gills and liver following exposure to waterborne indigo, benzo(a)pyrene and 3,3',4,4',5-pentachlorobiphenyl
2006 (English)In: Aquatic Toxicology, ISSN 0166-445X, E-ISSN 1879-1514, Vol. 79, no 3, p. 226-232Article in journal (Refereed) Published
Abstract [en]

We have developed a gill-filament based ethoxyresorufin O-deethylase (EROD) assay to be used as a tool to monitor cytochrome P4501A (CYP1A) induction in caged fish. The present study aimed to compare temporal patterns of EROD induction in gills and liver of rainbow trout (Oncorhynchus mykiss) exposed in the laboratory to readily metabolized and persistent CYP1A inducers, i.e. indigo, benzo[a]pyrene (BaP), and 3,3',4,4',5-pentachlorobiphenyl (PCB#126). Branchial and hepatic EROD activities were examined in fish exposed for 6, 12, or 24h and in fish exposed for 24h and then held in clean water for 2 or 14 days. Furthermore, branchial CYP1A protein expression was localized by immunohistochemistry. All compounds strongly induced branchial EROD activity within 6 h. The highest EROD inductions observed for indigo, BaP, and PCB#126 were roughly similar in gills (52-, 76-, and 74-fold), but differed considerably in liver (11-, 78-, and 200-fold). In indigo- and BaP-exposed fish, both hepatic and branchial EROD activities decreased rapidly in clean water. In PCB#126-exposed fish, decreased branchial and increased hepatic EROD activities were observed following transfer to clean water. The substances gave rise to immunostaining for CYP1A at different cellular sites. All inducers increased the CYP1A-immunostaining in the gill filament secondary lamellae, but PCB#126 also induced a pronounced CYP1A immunoreactivity in cells near the basal membrane of the epithelium of the primary lamellae. The observation that the low BaP and indigo concentrations induced EROD activity markedly in the gills but only slightly or not at all in the liver, supports the contention that readily metabolized AhR agonists may escape detection when hepatic EROD activity is used for environmental monitoring. The results show that gill filament EROD activity is a sensitive biomarker both for persistent and readily metabolized AhR agonists in polluted water.

Keywords
benzo[a]pyrene, CYP1A, gill, indigo, liver, 3, 3 ', 4, 4 ', 5-pentachlorobiphenyl (PCB#126)
National Category
Biological Sciences
Identifiers
urn:nbn:se:uu:diva-95921 (URN)10.1016/j.aquatox.2006.06.006 (DOI)000240567200003 ()16872689 (PubMedID)
Available from: 2007-05-09 Created: 2007-05-09 Last updated: 2017-12-14Bibliographically approved
3. Monitoring contaminants from oil production at sea by measuring gill EROD activity in Atlantic cod (Gadus morhua)
Open this publication in new window or tab >>Monitoring contaminants from oil production at sea by measuring gill EROD activity in Atlantic cod (Gadus morhua)
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2008 (English)In: Environmental Pollution, ISSN 0269-7491, E-ISSN 1873-6424, Vol. 153, no 1, p. 169-175Article in journal (Refereed) Published
Abstract [en]

An ex vivo gill EROD assay was applied in Atlantic cod (Gadus morhua) as a biomarker for waterborne CYP1A-inducing compounds derived from oil production at sea. Exposure to nominal concentrations of 1 ppm or 10 ppm North Sea crude oil in a static water system for 24 h caused a concentration-dependent gill EROD induction. Further, exposure of cod for 14 days to environmentally relevant concentrations of produced water (PW, diluted 1:200 or 1:1000) from a platform in the North Sea using a flow-through system resulted in a concentration-dependent induction of gill EROD. Crude oil (0.2 ppm) from the same oil field also proved to induce EROD. Finally, gill EROD activity in cod caged for 6 weeks at 500-10 000 m from two platforms outside Norway was measured. The activities in these fish were very low and did not differ from those in fish caged at reference sites.

Keywords
Atlantic cod, Biomarker, CYP1A, Crude oil, EROD, Gill, Produced water
National Category
Biological Sciences
Identifiers
urn:nbn:se:uu:diva-95922 (URN)10.1016/j.envpol.2007.07.025 (DOI)000255819300020 ()17854961 (PubMedID)
Available from: 2007-05-09 Created: 2007-05-09 Last updated: 2017-12-14Bibliographically approved
4. Gill EROD in monitoring of CYP1A inducers in fish: A study in rainbow trout (Oncorhynchus mykiss) caged in Stockholm and Uppsala waters
Open this publication in new window or tab >>Gill EROD in monitoring of CYP1A inducers in fish: A study in rainbow trout (Oncorhynchus mykiss) caged in Stockholm and Uppsala waters
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2007 (English)In: Aquatic Toxicology, ISSN 0166-445X, E-ISSN 1879-1514, Vol. 85, no 1, p. 1-8Article in journal (Refereed) Published
Abstract [en]

The gill filament 7-ethoxyresorufin O-deethylase (EROD) assay was evaluated as a monitoring tool for waterborne cytochrome P4501 A (CYP1A) inducers using rainbow trout (Oncorhynchus mykiss) caged in urban area waters in Sweden. To compare the CYP1A induction response in different tissues, EROD activity was also analyzed in liver and kidney microsomes. Immunohistochemistry was used to localize CYP1A protein in gill and kidney. In two separate experiments fish were caged at sites with fairly high expected polyaromatic hydrocarbon (PAH) contamination. In the first experiment, gill EROD activities were analyzed in fish exposed for 1-21 days in a river running through Uppsala. The reference site was upstream of Uppsala. In the second, gill, liver and kidney EROD activities were analyzed in fish exposed for 1-5 days in fresh or brackish waters of Stockholm and in a reference lake 60 km north of Stockholm. Fish exposed for 5 days followed by 2 days of recovery in tap water in the laboratory were also examined. The gill consistently showed a higher EROD induction compared with the liver and the kidney. After I day of caging, gill EROD activity was markedly induced (6-17-fold) at all sites examined. Induction in gill was pronounced (5-7-fold) also in fish caged at the reference sites. In the 21-day exposure study gill EROD activity remained highly induced throughout the experiment (26-fold at most) and the induced CYP1A protein was exclusively confined to the gill secondary lamellae. In the 5-day exposure experiment, EROD activity peaked after I day and then declined in both gill and liver, while CYP1A immunostaining in the gill remained intense over the 5-day period. In the kidney, CYP1A staining was weak or absent. We conclude that gill EROD activity is a more sensitive biomarker of exposure to waterborne CYP1A inducers than EROD activity in liver and kidney.

Keywords
fish; gill; CYP1A; EROD; monitoring
National Category
Biological Sciences
Identifiers
urn:nbn:se:uu:diva-95923 (URN)10.1016/j.aquatox.2007.07.013 (DOI)000250181300001 ()
Available from: 2007-05-09 Created: 2007-05-09 Last updated: 2017-12-14Bibliographically approved
5. Inhibition of CYP1A activity in fish detected by the gill filament EROD assay - studies on ketoconazole, bitertanol, acacetin and omeprazole
Open this publication in new window or tab >>Inhibition of CYP1A activity in fish detected by the gill filament EROD assay - studies on ketoconazole, bitertanol, acacetin and omeprazole
(English)Manuscript (Other (popular science, discussion, etc.))
Identifiers
urn:nbn:se:uu:diva-95924 (URN)
Available from: 2007-05-09 Created: 2007-05-09 Last updated: 2010-01-14Bibliographically approved

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