Abstract
Pharmaceuticals, such as anti-inflammatory nonsteroidal drugs, are frequently detected in aquatic ecosystems. Studies about the effects of these substances in nontarget organisms, such as bivalves, are relevant. The aim of this study was to evaluate the effects on antioxidant status caused by ibuprofen (IBU) in oysters Crassostrea gigas exposed for 1, 4, and 7 days at concentrations 1 and 100 μg L−1. Levels of IBU in tissues of oysters, as well as cell viability of hemocytes, were measured. The transcription of cytochrome P450 genes (CYP2AU2, CYP356A1, CYP3071A1, CYP30C1), glutathione S-transferase isoforms (GST-ω-like and GST-π-like), cyclooxygenase-like (COX-like), fatty acid binding protein-like (FABP-like), caspase-like, heat shock protein-like (HSP70-like), catalase-like (CAT-like), and the activity of catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR), and glutathione S-transferase (GST) were also evaluated in the gills of oysters. The highest levels of IBU were observed in animals exposed to 100 μg L−1. A significant upregulation of CYP2AU1, CYP356A1, CYP3071A1, GST-ω-like, GST-π-like, COX-like, and FABP-like was observed in oysters exposed to IBU under different experimental conditions. Oysters exposed to 1 μg L−1 for 7 days showed a significantly higher transcription of CYP2AU2, CYP356A1, CYP3071A1, GST-ω-like, and GST-π-like but lower GR activity. In conclusion, C. gigas exposed to environmentally relevant concentrations of IBU (1 μg L−1) exhibited increased transcription of certain genes and alterations on antioxidant and auxiliary enzymes, which could, in the the long term, cause damages to exposed organisms.
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References
Aebi H (1984) Catalase. In: Methods of enzymatic analysis. Academic press, London, pp 121–126
Ashton D et al (2004) Investigating the environmental transport of human pharmaceuticals to streams in the United Kingdom. Sci Total Environ 333:167–184. doi:10.1016/j.scitotenv.2004.04.062
Bendz D et al (2005) Occurrence and fate of pharmaceutically active compounds in the environment, a case study: Höje River in Sweden. J Hazard Mater 122:195–204. doi:10.1016/j.jhazmat.2005.03.012
Board PG (2011) The omega-class glutathione transferases: structure, function, and genetics. Drug Metab Rev 43:226–235. doi:10.3109/03602532.2011.561353
Boutet I et al (2004) Response of the Pacific oyster Crassostrea gigas to hydrocarbon contamination under experimental conditions. Gene 329:147–157. doi:10.1016/j.gene.2003.12.027
Bradford M (1976) A rapid and sensitive method for the quantification of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254. doi:10.1016/0003-2697(76)90527-3
Brun GL et al (2006) Pharmaceutically active compounds in Atlantic Canadian sewage treatment plant effluents and receiving waters, and potential for environmental effects as measured by acute and chronic aquatic toxicity. Environ Toxicol Chem 25:2163–2176
Calabrese EJ, Baldwin LA (2003) Hormesis: the dose-response revolution. Annu Rev Pharmacol Toxicol 43:175–197. doi:10.1146/annurev.pharmtox.43.100901.140223
Carlberg I, Mannervik B (1985) Glutathione reductase. Meth Enzymol 113:484–490. doi:10.1016/S0076-6879(85)13062-4
Contardo-Jara V et al (2011) Exposure to human pharmaceuticals carbamazepine, ibuprofen and bezafibrate causes molecular effects in Dreissena polymorpha. Aquat Toxicol 105:428–437. doi:10.1016/j.aquatox.2011.07.017
Daughton CG, Ternes TA (1999) Pharmaceuticals and personal care products in the environment: agents of subtle change? Environ Health Perspect 107:907–938
Ericson H, Thorsen G, Kumblad L (2010) Physiological effects of diclofenac, ibuprofen and propranolol on Baltic Sea blue mussels. Aquat Toxicol 99:223–231. doi:10.1016/j.aquatox.2010.04.017
Esteves A, Ehrlich R (2006) Invertebrate intracellular fatty acid binding proteins. Comp Biochem Physiol C 142:262–274. doi:10.1016/j.cbpc.2005.11.006
Farré ML et al (2001) Determination of drugs in surface water and wastewater samples by liquid chromatography-mass spectrometry: methods and preliminary results including toxicity studies with Vibrio fischeri. J Chromatogr A 938:187–197
Fent K, Weston AA, Caminada D (2006) Ecotoxicology of human pharmaceuticals. Aquat Toxicol 76(2):122–159. doi:10.1016/j.aquatox.2005.09.009
Ferrando-Climent L et al (2012) Comprehensive study of ibuprofen and its metabolites in activated sludge batch experiments and aquatic environment. Sci Total Environ 438:404–413. doi:10.1016/j.scitotenv.2012.08.073
Gagné F et al (2005) Inflammatory properties of municipal effluents to Elliptio complanata mussels—lack of effects from anti-inflammatory drugs. Comp Biochem Physiol C 141:332–337. doi:10.1016/j.cca.2005.06.006
García-Martín E et al (2004) Interindividual variability in ibuprofen pharmacokinetics is related to interaction of cytochrome P450 2C8 and 2C9 amino acid polymorphisms. Clin Pharmacol Therap 76:119–127. doi:10.1016/j.clpt.2004.04.006
Gierse JK et al (1999) Kinetic basis for selective inhibition of cyclo-oxygenases. Biochem J 339:607–614
Gómez M et al (2007) Pilot survey monitoring pharmaceuticals and related compounds in a sewage treatment plant located on the Mediterranean coast. Chemosphere 66:993–1002. doi:10.1016/j.chemosphere.2006.07.051
Gonzalez-Rey M, Bebianno MJ (2011) Non-steroidal anti-inflammatory drug (NSAID) ibuprofen distresses antioxidant defense system in mussel Mytilus galloprovincialis gills. Aquat Toxicol 105:264–269. doi:10.1016/j.aquatox.2011.06.015
Gonzalez-Rey M, Bebianno MJ (2012) Does non-steroidal antiinflammatory (NSAID) ibuprofen induce antioxidant stress and endocrine disruption in mussel Mytilus galloprovincialis? Environ Toxicol Pharmacol 33:361–371. doi:10.1016/j.etap.2011.12.017
Hamman MA et al (1997) Regioselective and stereoselective metabolism of ibuprofen by human cytochrome P450 2C. Biochem Pharmacol 54:33–41. doi:10.1371/journal.pone. 0012329
Hermes-Lima M (2004) Oxygen in biology and biochemistry: role of free radicals. In: Storey KB (ed) Functional metabolism: regulation and adaptation. Wiley, New Jersey, pp 319–368
Hernando MD et al (2006) Environmental risk assessment of pharmaceutical residues in wastewater effluents, surface waters and sediments. Talanta 69:334–342. doi:10.1016/j.talanta.2005.09.037
Huerta B et al (2012) Pharmaceuticals in biota in the aquatic environment: analytical methods and environmental implications. Anal Bioanal Chem 404:2611–2624. doi:10.1007/s00216-012-6144-y
IMS Institute for Healthcare informatics (2011). The global use of medicines: outlook through 2015. pp. 27.http://www.imshealth.com/deployedfiles/ims/Global/Content/Insights/IMS%20Institute%20for%20Healthcare%20Informatics/Global_Use_of_Medicines_Report.pdf
Keen JH, Habig WH, Jakoby W (1976) Mechanism for the several activities of the glutathione S-transferase. J Biol Chem 251:6183–6188
Kim SD et al (2007) Occurrence and removal of pharmaceuticals and endocrine disruptors in South Korean surface, drinking, and waste waters. Water Res 41:1013–1021. doi:10.1016/j.watres.2006.06.034
Kümmerer K (2009) The presence of pharmaceuticals in the environment due to human use—present knowledge and future challenges. J Environ Manag 90:2354–2366. doi:10.1016/j.jenvman.2009.01.023
Leemann TD et al (1993) A major role for cytochrome P450TB (CYP2C subfamily) in the actions of non-steroidal antiinflammatory drugs. Drugs Exp Clin Res 19:189–195. doi:10.1371/journal.pone.0012329
Lüchmann KH et al (2014) A light in the darkness: new biotransformation genes, antioxidant parameters and tissue-specific responses in oysters exposed to phenanthrene. Aquat Toxicol 152:324–334. doi:10.1016/j.aquatox.2014.04.021
Madden JC et al (2009) Pharmaceuticals in the environment: good practice in predicting acute ecotoxicological effects. Toxicol Lett 185:85–101. doi:10.1016/j.toxlet.2008.12.005
Manduzio H et al (2004) Seasonal variations in antioxidant defences in blue mussels Mytilus edulis collected from a polluted area: major contributions in gills of an inducible isoform of Cu/Zn-superoxide dismutase and of glutathione S-transferase. Aquat Toxicol 70:83–93. doi:10.1016/j.aquatox.2004.07.003
Markert A et al (2010) Recently established Crassostrea-reefs versus native Mytilus-beds: differences in ecosystem engineering affects the macrofaunal communities (Wadden Sea of Lower Saxony, southern German Bight). Biol Invasions 12:15–32. doi:10.1007/s10530-009-9425-4
Matozzo V et al (2012) The nonsteroidal anti-inflammatory drug, ibuprofen, affects the immune parameters in the clam Ruditapes philippinarum. Mar Environ Res 79:116–121. doi:10.1016/j.marenvres.2012.06.003
Medeiros ID et al (2008a) Differential gene expression in oyster exposed to sewage. Mar Environ Res 66:156–157. doi:10.1016/j.marenvres.2008.02.048
Medeiros ID et al (2008b) Induced gene expression in oyster Crassostrea gigas exposed to sewage. Environ Toxicol Pharmacol 26:362–365. doi:10.1016/j.etap.2008.05.004
Milan M et al (2013) Gene transcription and biomarker responses in the clam Ruditapes philippinarum after exposure to ibuprofen. Aquat Toxicol 126:17–29. doi:10.1016/j.aquatox.2012.10.007
Osada M, Nomura T (1990) The levels of reproductive cycle prostaglandins associated with the reproductive cycle of the scallop, Patinopecten yessoensis. Prostaglandins 40(3):229–239. doi:10.1016/0090-6980(90)90011-J
Parolini M et al (2009) An in vitro biomarker approach for the evaluation of the ecotoxicity of non-steroidal anti-inflammatory drugs (NSAIDs). Toxicol In Vitro 23:935–942. doi:10.1016/j.tiv.2009.04.014
Parolini M et al (2011) Chronic effects induced by ibuprofen on the freshwater bivalve Dreissena polymorpha. Ecotox Environ Safe 74:1586–1594. doi:10.1016/j.ecoenv.2011.04.025
Pounds N et al (2008) Acute and chronic effects of ibuprofen in the mollusc Planorbis carinatus (Gastropoda: Planorbidae). Ecotox Environ Safe 70:47–52. doi:10.1016/j.ecoenv.2007.07.003
Qiagen (2009a). QIAzol handbook: for efficient lysis of fatty tissues and all other types of tissue before RNA purification
Qiagen (2009b). QuantiTect® reverse transcription handbook, pp 31
Qiagen (2011). QuantiFast® SYBR® Green PCR KIT, Quick Start Protocol, pp 3
Rao P, Knaus EE (2008) Evolution of nonsteroidal anti-inflammatory cyclooxygenase (COX) inhibition and beyond drugs (NSAIDs). J Pharm Pharmaceut Sci 11(2):81s–110s
Regoli F, Principato G (1995) Glutathione, glutathione-dependent and antioxidant enzymes in mussel, Mytilus galloprovincialis, exposed to metals under field and laboratory conditions: implications for the use of biochemical biomarkers. Aquat Toxicol 31:143–164. doi:10.1016/0166-445X(94)00064-W
Regoli F et al (2002) Oxidative stress in ecotoxicology: from analysis of individual antioxidants to a more integrated approach. Mar Environ Res 54:419–423. doi:10.1016/S0141-1136(02)00146
Regoli F et al (2011) Molecular and biochemical biomarkers in environmental biomonitoring: a comparison of biotransformation and antioxidant defense systems in multiple tissues. Aquat Toxicol 105:56–66. doi:10.1016/j.aquatox.2011.06.014
Santos LH et al (2010) Ecotoxicological aspects related to the presence of pharmaceuticals in the aquatic environment. J Hazard Mater 175(1–3):45–95. doi:10.1016/j.jhazmat.2009.10.100
Ternes TA, Joss A, Siegrist H (2004) Scrutinizing pharmaceuticals and personal care products in wastewater treatment. Environ Sci Technol 38(20):392A–399A. doi:10.1021/es040639t
Toledo-Silva G et al (2008) Cloning a new cytochrome P450 isoform (CYP356A1) from oyster Crassostrea gigas. Mar Environ Res 66:15–18. doi:10.1016/j.marenvres.2008.02.010
Tong AY, Peake PM, Braund R (2011) Disposal practices for unused medications around the world. Environ Int 37:292–298. doi:10.1016/j.envint.2010.10.002
Valavanidis A et al (2006) Molecular biomarkers of oxidative stress in aquatic organisms in relation to toxic environmental pollutants. Ecotox Environ Safe 64:178–189. doi:10.1016/j.ecoenv.2005.03.013
Verenitch SS, Lowe CJ, Mazumder A (2006) Determination of acidic drugs and caffeine in municipal wastewaters and receiving waters by gas chromatography ion trap tandem mass spectrometry. J Chromatogr A 1116:193–203. doi:10.1016/j.chroma.2006.03.005
Weigel S et al (2004) Determination of selected pharmaceuticals and caffeine in sewage and seawater from Tromsø/Norway with emphasis on ibuprofen and its metabolites. Chemosphere 56:583–592. doi:10.1016/j.chemosphere.2004.04.015
Wendel A (1981) Glutathione peroxidase. Meth Enzymol 77:325–333. doi:10.1016/S0076-6879(81)77046-0
Wright JM et al (2014) Populations of Pacific oysters Crassostrea gigas respond variably to elevated CO2 and predation by Morula marginalba. Biol Bull 226:269–281
Zar JH (1999) Biostatistical analysis. Prentice-Hall, Englewood Cliffs
Zhang G et al (2013) The oyster genome reveals stress adaptation and complexity of shell formation. Nature 490:49–54. doi:10.1038/nature11413
Zuccato E et al (2005) Identification of the pharmaceuticals for human use contaminating the Italian aquatic environment. J Hazard Mater 122:205–209. doi:10.1016/j.jhazmat.2005.03.001
Acknowledgments
This research was partially funded by the project UNIVERSAL - MCTI / CNPq No. 14/2012 (483028/2012-6). Miguel Angel Saldaña Serrano was a fellow master’s of the “Programa de Estudantes - Convenio de Pós-Graduação” (PEC-PG/2011)/Conselho Nacional de Desenvolvimento Cientifico e Tecnológico (CNPq). Dr. Afonso Celso Dias Bainy and Dr. Cláudio Manoel Rodrigues de Melo are recipients of the CNPq productivity fellowship. Maria João Bebianno is a recipient of the CNPq PVE fellowship. The authors would like to thank the LMM (UFSC) for the donation of oysters used in this work and the LCM (UFSC) for preparing the samples sent for IBU analysis. The research leading to these results was partly funded by the EU project GENERA within the framework of the Marie Curie IRSES Actions (FP7-PEOPLE-2009-IRSES) Proposal no. 247559.
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Supplementary Figure 1
Cellular viability of hemocytes (mean ± standard deviation (SD)) was measured in hemolymph of oysters Crassostrea gigas exposed to IBU 0, 1 and 100 μg.L−1 for 1, 4 and 7 days. Sample size (n) varied from 8 to 10 individuals in each group. (PDF 15 kb)
Supplementary Figure 2
Caspase-like and HSP 70-like gene transcription profiles after 1 day (a, d), 4 days (b, e) and 7 days (c, f) of exposure to IBU 1 and 100 μg.L−1. Same letters indicate not significant differences (p > 0.05) (PDF 39 kb)
Supplementary Figure 3
CYP30C1-like and catalase-like (CAT-like) gene transcription profiles after 1 day (a, d), 4 days (b, e) and 7 days (c, f) of exposure to IBU 1 and 100 μg.L−1. Same letters indicate not significant differences (p > 0.05) (PDF 38 kb)
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Serrano, M.A.S., Gonzalez-Rey, M., Mattos, J.J. et al. Differential gene transcription, biochemical responses, and cytotoxicity assessment in Pacific oyster Crassostrea gigas exposed to ibuprofen. Environ Sci Pollut Res 22, 17375–17385 (2015). https://doi.org/10.1007/s11356-014-4023-0
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DOI: https://doi.org/10.1007/s11356-014-4023-0