Skip to main content
SpringerLink
Log in

Oxidative stress in liver cell culture from mullet, Liza klunzingeri, induced by short-term exposure to benzo[a]pyrene and nonylphenol

  • Published:
Fish Physiology and Biochemistry Aims and scope Submit manuscript

Abstract

The present investigation aimed to use primary liver cell culture obtained from mullet, Liza klunzingeri, to evaluate the toxic effects of benzo[a]pyrene (BaP) and nonylphenol (NP) on the antioxidant defense system. Liver samples taken from 20 L. klunzingeri were digested with 0.1% collagenase IV. The digested cells were then moved to Leibovitz L-15 culture medium and incubated at 25 °C for 2 weeks. 10−5 mol/l of BaP and 10−4 mol/l of NP were considered as the half maximal inhibitory concentration (IC50). Cells were then incubated with L-15 medium containing BaP (0[control], 10−6,2 × 10−6,3 × 10−6 mol/l) and NP (0[control],10−5,2 × 10−5,3 × 10−5 mol/l), and sampling was performed after 6, 12, and 24 h of incubation for measurement of catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), lipid peroxidation (LPO), total antioxidant power, and total protein. The lowest concentration of BaP and NP did not have considerable toxic effects on cultivated hepatocytes. The activities of SOD, CAT, GPx, LPO, total antioxidant power, and total protein changed dose-dependently in cells treated with BaP and NP. In conclusion, based on the results, short-term exposure to BaP and NP induced the oxidative stress in cultivated liver cells of L. klunzingeri. The toxicity of both pollutants is mainly because of the induction of the reactive oxygen species (ROS), which lead to cell membrane disruption, damage of cellular metabolism, and interference with cellular macromolecules.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  • Abdul Majeed S, Nambi KSN, Taju G, Sundar Raj N, Madan N, Sahul Hameed AS (2013) Establishment and characterization of permanent cell line from gill tissues of Labeo rohita (Hamilton) and its application in gene expression and toxicology. Cell Biol Toxicol 29:59–73

    CAS  PubMed  Google Scholar 

  • Adachi T, Yasuda K, Mori C, Yoshinaga M, Aoki N, Tsujimoto G (2005) Promoting insulin secretion in pancreatic islets by means of biphenol A and nonylphenol via intracellular estrogen receptors. Food Chem Toxicol 43(5):713–719

    CAS  PubMed  Google Scholar 

  • Aly HA, Domènech O, Banjar ZM (2012) Effect of nonylphenol on male reproduction: analysis of rat epididymal biochemical markers and antioxidant defense enzymes. Toxicol Appl Pharmacol 261:134–141

    CAS  PubMed  Google Scholar 

  • Aoki M, Kurasaki M, Saito T, Seki S, Hosokawa T, Takahashi Y, Fujita H, Iwakuma T (2004) Nonylphenol enhances apoptosis induced by serum deprivation in PC12 cells. Life Sci 74(18):2301–2312

  • ATSDR (Agency for Toxic Substances and Disease Registry) (1995) Toxicological profile for polycyclic aromatic hydrocarbons (PAHs)

  • Babich H, Borenfreund E (1991) Cytotoxicity and genotoxicity assays with cultured fish cells: a review. Toxicol in Vitro 5:91–100

    CAS  PubMed  Google Scholar 

  • Banaee M, Sureda A, Mirvaghefi A, Ahmadi K (2011) Effects of diazon on biochemical parameters of blood in rainbow trout (Oncorhynchus mykiss). Pestic Biochem Physiol 99:1–6

    CAS  Google Scholar 

  • Benzie IF, Strain JJ (1999) Ferric reducing/antioxidant power assay: direct measure of total antioxidant activity of biological fluids and modified version for simultaneous measurement of total antioxidant power and ascorbic acid concentration. Methods Enzymol 299:15–27

    CAS  PubMed  Google Scholar 

  • Borenfreund E, Puerner JA (1985) Toxicity determined in vitro by morphological alterations and neutral red absorption. Toxicol Lett 24:119–124

    CAS  PubMed  Google Scholar 

  • Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Reproduction Research Laboratories, Department of Biochemistry. University of Georgia. Anal Biochem 72:248–254

    CAS  PubMed  Google Scholar 

  • Briede JJ, Godschalk RW, Emans MT, De Kok TM, Van Agen E, Van Maanen J, Van Schooten FJ, Kleinjans JC (2004) In vitro and in vivo studies on oxygen free radical and DNA adduct formation in rat lung and liver during benzo[a]pyrene metabolism. Free Radic Res 38:995–1002

    CAS  PubMed  Google Scholar 

  • Castano A, Bols N, Braunbeck T, Dierickx P, Halder M, Isomaa B, Kawahara K, Lee LE, Mothersill C, Part P, Repetto G, Sintes JR, Rufli H, Smith R, Wood C, Senger H (2003) The use of fish cells in ecotoxicology. ATLA 31(3):317–351

    CAS  PubMed  Google Scholar 

  • Cui Q, Chen F, Chen H, Peng H, Wang K (2019) Benzo[a]pyrene (BaP) exposure generates persistent reactive oxygenspecies (ROS) to inhibit the NF-kB pathway in medaka (Oryziasmelastigma). Environ Poll 251:502–509

    CAS  Google Scholar 

  • De Lamirande E, Gagnon C (1992) Reactive oxygen species and human spermatozoa. I Effects on the motility of intact spermatozoa and on sperm axonemes. J Androl 13:368–378

    PubMed  Google Scholar 

  • Di Giulio RT (1991) Indices of oxidative stress as biomarkers for environmental contamination. In: Mayes MA, Barron MG (eds) Aquatic toxicology and risk assessment. American Society for Testing and Materials, Philadelphia, pp 15–31

    Google Scholar 

  • Di Giulio RT, Hinton DE (2008) The toxicology of fishes. Taylor and Francis Group, Boca Raton

    Google Scholar 

  • Di Giulio RT, Washburn PC, Wenning RJ (1989) Biochemical responses in aquatic animals: a review of determinants of oxidative stress. Environ Toxicol Chem 8:1103–1123

    Google Scholar 

  • Doba T, Burton GW, Ingold KU (1985) Antioxidant and co-antioxidant activity of vitamin C. The effect of vitamin C, either alone or in the presence of vitamin E or a water-soluble vitamin E analogue, upon the peroxidation of aqueous multilamellar phospholipid liposomes. Biochim Biophys Acta 835(2):298–303

    CAS  PubMed  Google Scholar 

  • Eberhart J, Coffing SL, Anderson JN, Marcus C, Kalogeris TJ, Baird WM, Park SS, Gelboin HV (1992) The time-dependent increase in the binding of benzo[a]pyrene to DNA through (+)-anti benzo[a]pyrene 7,8-diol-9,10-epoxide in primary rat hepatocyte cultures results from induction of cytochrom P4501A1 by benzo[a]pyrene treatment carcinogenesis. Carcinogenesis 13(2):297–301

    CAS  PubMed  Google Scholar 

  • Fent K (2001) Fish cell lines as versatile tools in ecotoxicology: assessment of cytotoxicity, cytochrome P4501A induction potential and estrogenic activity of chemicals and environmental samples. Toxicol in Vitro 15:477–488

    CAS  PubMed  Google Scholar 

  • Frouin H, Pellerin J, Fournier M, Pelletier E, Richard P, Pichaud N, Rouleau C, Garnerot F (2007) Physiological effects of polycyclic aromatic hydrocarbonson soft-shell clam Mya arenaria. Aquat Toxicol 82:120–134

    CAS  PubMed  Google Scholar 

  • Ganie SA, Haq E, Hamid A, Masood A, Zargar MA (2011) Long dose exposure of hydrogen peroxide (H2O2) in albino rats and effect of Podaphyllum hexandrum on oxidative stress. Eur Rev Med Pharmacol Sci 15:906–915

    CAS  PubMed  Google Scholar 

  • Gasparovic AC, Jaganjac M, Mihaljevic B, Sunjic SB, Zarkovic N (2013) Assays for the measurement of lipid peroxidation. Methods Mol Biol 965:283–296

    CAS  PubMed  Google Scholar 

  • Gate L, Paul J, Ba GN, Tew KD, Tapiero H (1999) Oxidative stress induced in pathologies: the role of antioxidants. Biomed Pharmacother 53:169–180

    CAS  PubMed  Google Scholar 

  • Halliwell B (1994) Free radicals, antioxidants, and human disease: curiosity, cause, or consequence? Lancet 344:721–724

    CAS  PubMed  Google Scholar 

  • Hamdy RD, Cornelis G, Fernandes T, Tsyusko O, Decho A, Sabo-Ahwood T, Metcalfe C, Steevens JA, Klains SJ, Koelmans AA, Horne N (2012) Ecotoxicity test methods for engineered nanomaterials: practical experience and recommendations from the bench. Environ Toxicol Chem 31:15–31

    Google Scholar 

  • Hughes PJ, McLellan H, Lowes DA, Zafar Khan S, Bilmen JG, Tovey SC (2000) Estrogenic alkylphenols induce cell death by inhibiting testis endoplasmic reticulum Ca2+ pumps. Biochem Bioph Res Co 277:568–574

    CAS  Google Scholar 

  • Jifa W, Zhiming Y, Xiuxian S, You W (2006) Response of integrated biomarkers of fish (Lateolabrax japonicus) exposed to benzo(a)pyrene and sodium dodecylbenzene sulfonate. Ecotoxicol Environ Saf 65:223–236

    Google Scholar 

  • Joseph P, Jaiswal AK (1998) NAD(P)H:quinone oxidoreductase 1 reduces the mutagenicity of DNA caused by NADPH:P450 reductase-activated metabolites of benzo(a)pyrene quinones. Br J Cancer 77:709–719

    CAS  PubMed  PubMed Central  Google Scholar 

  • Kazeto Y, Place AR, Trant JM (2004) Effects of endocrine disrupting chemicals on the expression of CYP19 genes in zebrafish (Danio rerio) juveniles. Aquat Toxicol 69:25–34

    CAS  PubMed  Google Scholar 

  • Khaniyan M, Salamat N, Safahieh A, Movahedinia A (2014) Detection of benzo[a]pyrene-induced immunotoxicity in orange spotted grouper (Epinephelus coioides). Environ Toxicol. https://doi.org/10.1002/tox.22047

  • Kim KB, Lee BM (1997) Oxidative stress to DNA, protein, and antioxidant enzymes (superoxide dismutase and catalase) in rats treated with benzo(a)pyrene. Cancer Lett 113:205–212

    CAS  PubMed  Google Scholar 

  • Kim BH, Takemura A (2003) Culture conditions affect induction of vitelligenin synthesis by estradiol-17b in primary cultures of tilapia hepatocytes. Comp Biochem Physiol B 135:231–239

    CAS  PubMed  Google Scholar 

  • Kudo C, Wada K, Masuda T, Yonemura T, Shibuya A, Fujimoto Y (2004) Nonylphenol induces the death of neural stem cells due to activation of the caspase cascade and regulation of the cell cycle. J Neurochem 88(6):1416–1423

    CAS  PubMed  Google Scholar 

  • Liang Y, Wong CKC, Xu Y, Wong MH (2004) Effects of 2,3,7,8-TCDD and benzo[a]pyrene on modulating vitellogenin expression in primary culture of crucian carp (Carassius auratus) hepatocytes. Chin Sci Bull 49:2372–2378

    CAS  Google Scholar 

  • Livingstone MB, Prentice AM, Strain JJ, Coward WA, Black AE, Barker ME (1990) Accuracy of weighed dietary records in studies of diet and health. Br Med J 300:708–712

    CAS  Google Scholar 

  • Maes M, Fisar Z, Medina M, Scapagnini G (2012) New drug targets in depression: inflammatory, cell-mediated immune, oxidative and nitrosative stress, mitochondrial, antioxidant, and neuroprogressive pathways and new drug candidates-NrF2 activators and GSK-3 inhibitors. Inflammopharmacol 20(3):127–150

    CAS  Google Scholar 

  • Mahdinezhad Gorji N, Jorsaraei GH, Hojati V, Zabihi E, Khalilpour A, Abedian Z, Tahmasbpour E (2015) A comparison between the cytotoxicity induced by gossypol in two testicular cell lines. Iran J Toxicol 8(27):1188–1195

    CAS  Google Scholar 

  • Mao Z, Zheng YL, Zhang YQ (2011) Behavioral impairment and oxidative damage induced by chronic application of nonylphenol. Int J Mol Sci 12(1):114–127

    CAS  Google Scholar 

  • Mao XY, Zhou HH, Jin WL (2019) Redox-Rrelated neuronal death and crosstalk as drug targets: focus on epilepsy. Front Neurosci 13:512

  • Midhila EM, Chitra KC (2015) Nonylphenol-induced hepatotoxicity in the freshwater fish, Oreochromis mossambicus. Int J Sci Res Publ 5(3):1–5

    Google Scholar 

  • Miller NJ, Rice-Evans C, Davies MJ, Gopinathan V, Milner A (1993) A novel method for measuring antioxidant capacity and its application to monitoring the antioxidant status in premature neonates. Clin Sci 84:407–419

    CAS  PubMed  Google Scholar 

  • Minatoya M, Itoh S, Yamazaki K, Araki A, Miyashita C, Tamura N, Yamamoto J, Onoda Y, Ogasawara K, Matsumura T, Kishi R (2018) Prenatal exposure to bisphenol a and phthalates and behavioral problems in children at preschool age: the Hokkaido study on environment and Children's health. Environ Health Prev Med 23:43

    PubMed  PubMed Central  Google Scholar 

  • Monteiro DA, Rantin FT, Kalinin AL (2009) The effects of selenium on oxidative stress biomarkers in the freshwater characid fish matrinxa (Brycon cephhalus) (Gunther, 1869) exposed to organophosphate insecticide Folisuper 600BR (methyl parathion). Comp Biochem Physiol C 149:40–49

    Google Scholar 

  • Mouithys-Mickalad A, Mareque-Faez J, Chistiaens L, Kohnen S, Deby C, Hoebeke M, Lamy M, Deby-Dupont G (2004) In vitro evaluation of glutathione peroxidase (GPx)-like activity and antioxidant properties of some Ebselen analogues. Redox Rep 9(2):81–87

    Google Scholar 

  • Newbold RR, Padilla-Banks E, Jefferson WN (2006) Adverse effects of the model environmental estrogen diethylstilbestrol are transmitted to subsequent generations. Endocrinol 147(6):S11–S17

    CAS  Google Scholar 

  • Oguz AR, Unal G (2011) The effects of 17α-ethynylestradiol, 4-nonylphenol and phenol red on vitellogenin synthesis in juvenile Chalcalburnus tarichi primary hepatocyte culture. Toxicol Ind Health 27(4):379–384

    CAS  Google Scholar 

  • Pan LQ, Ren J, Liu J (2006) Responses of antioxidant systems and LPO level to benzo[a]pyrene and benzo[k]flurantheren in the haemolymph of the scallop Chlamys ferrari. Environ Pollut 141:443–451

    CAS  PubMed  Google Scholar 

  • Pereira B, Rosa LF, Safi DA, Bechara EJ, Curi R (1995) Hormonal regulation of superoxide dismutase, catalase and glutathione peroxidase activities in rat macrophages. Biochem Pharmacol 50:2093–2098

    CAS  PubMed  Google Scholar 

  • Puerto M, Pichardo S, Jos A, Camean AM (2009) Oxidative stress induced by microcystin-LR on PLHC-1 fish cell line. Toxicol in Vitro 23:1445–1449

    CAS  PubMed  Google Scholar 

  • Rao JV (2006) Toxic effects of novel organophosphorus insecticide (RPR-V) on certain biochemical parameters of euryhaline fish, Oreochromis mossambicus. Pestic Biochem Physiol 86:78–84

    CAS  Google Scholar 

  • Rodd AL, Messier NJ, Vaslet CA, Kane AB (2017) A 3D fish liver model for aquatic toxicology: morphological changes and Cyp1a induction in PLHC-1 microtissues after repeated benzo(a)pyrene exposures. Aquat Toxicol 186:134–144

    CAS  PubMed  PubMed Central  Google Scholar 

  • Sandstrom J, Nilsson P, Karlsson K (1994) 10-fold increase in human plasma extracellular superoxide dismutase content caused by a mutation in heparin binding domain. J Biol Chem 269(29):19163–19166

    CAS  PubMed  Google Scholar 

  • Schimer K (2006) Proposal to improve vertebrate cell cultures to establish them as substitutes for the regulatory testing of chemicals and effluents using fish. Txicol 224:163–183

    Google Scholar 

  • Schug TT, Janesick A, Blumberg B, Heindel JJ (2011) Endocrine disrupting chemicals and disease susceptibility. J Steroid Biochem Mol Biol 127(3–5):204–215

    CAS  PubMed  PubMed Central  Google Scholar 

  • Soldatow VY, LeCluyse EL, Griffith LG, Rusyn I (2013) In vitro models for liver toxicity testing. Toxicol Res 2:23–29

    CAS  Google Scholar 

  • Sole M, Porte C, Albaige SL (1995) The use of biomarkers for assessing the effects of organic pollution in mussels. Sci Total Environ 159:147–153

    CAS  PubMed  Google Scholar 

  • Taju G, Abdul Majeed S, Nambi KSN, Sahul Hameed AS (2014) In vitro assay for the toxicity of silver nanoparticles using heart and gill cell lines of Catla catla and gill cell line of Labeo rohita. Comp Biochem Physiol. https://doi.org/10.1016/j.cbpc.2014.01.007

  • Thannickal VJ, Fanburg BL (2000) Reactive oxygen species in cell signaling. Am Physiol Soc Lung Cell Mol Physiol 279(6):1005–1028

    Google Scholar 

  • Thomas MG, Lawson C, Allanson NM, Leslie BW, Bottomley JR, McBride A, Olusanua OA (2003) A series of 2(z)-2-benzylidene-6,7-dihydrobenzofuran-3[2H]-ones as inhibitors of chorismate synthase. Bioorg Med Chem Lett 13:423–426

    CAS  PubMed  Google Scholar 

  • US-EPA (U.S. Environmental Protection Agency) (1990) Guidance for Data Usability in Risk Assessment. U.S. Environmental Protection Agency, Washington, DC. EPA/540/G-90/008

  • Wei W, Li X, Li XN, Chen X, Liu A, Lu W (2010) Oxidative stress and cell-cycle change induced by coexposed PCB126 and benzo(a) pyrene to human hepatoma (HepG2) cells. Environ Toxicol DOI. https://doi.org/10.1002/tox.20649

  • Wen CM, Lee CW, Wang CS, Cheng YH, Huang HY (2008) Development of two cell lines from Epinephelus coioides brain tissue for characterization of betanodavirus and megalocytivirus infectivity and propagation. Aquaculture 278:14–21

    CAS  Google Scholar 

  • Weydert CJ, Cullen J (2010) Measurement of superoxide dismutase, catalase, and glutathione peroxidase in cultured cells and tissue. Nat Protoc 5(1):51–66

    CAS  PubMed  Google Scholar 

  • Wu M, Xu H, Shen Y, Qiu W, Yang M (2011) Oxidative stress in zebrafish embryos induced by short-term exposure to bisphonel A, nonylphenol, and their mixture. Environ Toxicol Chem 30(10):2335–2341

    CAS  PubMed  Google Scholar 

  • Zacchino V, Centoducati G, Narracci M, Selvaggi M, Santacroce MP (2013) Effect of Benzo[a]Pyrene on Gilthead Sea bream (Sparus aurata L.) hepatocyte exposed in vitro to short and long trials. Ital J Anim Sci 12:1–17

    Google Scholar 

  • Zhou B, Liu C, Wang J, Lam PK, Wu RS (2006) Primary cultured cells as sensitive in vitro model for assessment of toxicants-comparison to hepatocytes and gill epithelia. Aquat Toxicol 80(2):109–118

    CAS  PubMed  Google Scholar 

Download references

Funding

This work was supported by the Khorramshahr University of Marine Science and Technology [grant number 167].

Author information

Authors and Affiliations

  1. Department of Marine Biology, Faculty of Marine Sciences, Khorramshahr University of Marine Science and Technology, Khorramshahr, Iran

    Negin Salamat & Negin Derakhshesh

Authors
  1. Negin Salamat
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Negin Derakhshesh
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Negin Salamat.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Salamat, N., Derakhshesh, N. Oxidative stress in liver cell culture from mullet, Liza klunzingeri, induced by short-term exposure to benzo[a]pyrene and nonylphenol. Fish Physiol Biochem 46, 1183–1197 (2020). https://doi.org/10.1007/s10695-020-00783-y

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10695-020-00783-y

Keywords

Search

Navigation