Methyl mercury (MeHg) is an organic highly toxic compound that is transported efficiently via the human placenta. Our previous data suggest that MeHg is taken up into placental cells by amino acid transporters while mercury export from placental cells mainly involves ATP binding cassette (ABC) transporters. We hypothesized that the ABC transporter multidrug resistance-associated protein (MRP)1 (ABCC1) plays an essential role in mercury export from the human placenta. Transwell transport studies with MRP1-overexpressing Madin-Darby Canine Kidney (MDCK)II cells confirmed the function of MRP1 in polarized mercury efflux. Consistent with this, siRNA-mediated MRP1 gene knockdown in the human placental cell line HTR-8/SVneo resulted in intracellular mercury accumulation, which was associated with reduced cell viability, accompanied by increased cytotoxicity, apoptosis, and oxidative stress as determined via the glutathione (GSH) status. In addition, the many sources claiming different localization of MRP1 in the placenta required a re-evaluation of its localization in placental tissue sections by immunofluorescence microscopy using an MRP1-specific antibody that was validated in-house. Taken together, our results show that (1) MRP1 preferentially mediates apical-to-basolateral mercury transport in epithelial cells, (2) MRP1 regulates the GSH status of placental cells, (3) MRP1 function has a decisive influence on the viability of placental cells exposed to low MeHg concentrations, and (4) the in situ localization of MRP1 corresponds to mercury transport from maternal circulation to the placenta and fetus. We conclude that MRP1 protects placental cells from MeHg-induced oxidative stress by exporting the toxic metal and by maintaining the placental cells' GSH status in equilibrium.

Clinic for Pediatrics and Adolescent Medicine University Hospital Tulln Tulln Austria

Department of Obstetrics and Gynecology Medical University of Graz Graz Austria

Department of Obstetrics and Gynecology Medical University Vienna Vienna Austria

Department of Pharmacology and Toxicology Charles University Hradec Kralove Czech Republic

Institute of Medical Genetics Medical University of Vienna Vienna Austria

Institute of Pathophysiology and Allergy Research Medical University of Vienna Vienna Austria

Karl Landsteiner Private University for Health Sciences Krems Austria

See more in PubMed

Afrouzian M, Al-Lahham R, Patrikeeva S, et al. Role of the efflux transporters BCRP and MRP1 in human placental bio-disposition of pravastatin. Biochem Pharmacol. 2018;156:467–478. doi: 10.1016/j.bcp.2018.09.012. PubMed DOI PMC

Andreoli V, Sprovieri F. Genetic aspects of susceptibility to mercury toxicity: an overview. Int J Environ Res Public Health. 2017;14(1):93. doi: 10.3390/ijerph14010093. PubMed DOI PMC

Atkinson DE, Greenwood SL, Sibley CP, Glazier JD, Fairbairn LJ. Role of MDR1 and MRP1 in trophoblast cells, elucidated using retroviral gene transfer. Am J Physiol Cell Physiol. 2003;285(3):C584–C591. doi: 10.1152/ajpcell.00418.2002. PubMed DOI

Ballatori N, Krance SM, Marchan R, Hammond CL. Plasma membrane glutathione transporters and their roles in cell physiology and pathophysiology. Mol Aspects Med. 2009;30(1–2):13–28. doi: 10.1016/j.mam.2008.08.004. PubMed DOI PMC

Balthasar C, Stangl H, Widhalm R, Granitzer S, Hengstschlager M, Gundacker C. Methylmercury Uptake into BeWo cells depends on LAT2–4F2hc, a system L amino acid transporter. Int J Mol Sci. 2017;18(8):1730. doi: 10.3390/ijms18081730. PubMed DOI PMC

Benirschke K, Burton G, Baegen R. Pathology of the human placenta. Berlin: Springer; 2012.

Biondi C, Ferretti ME, Lunghi L, et al. cAMP efflux from human trophoblast cell lines: a role for multidrug resistance protein (MRP)1 transporter. Mol Hum Reprod. 2010;16(7):481–491. doi: 10.1093/molehr/gaq023. PubMed DOI

Blaschitz A, Gauster M, Dohr G. Application of cryo-compatible antibodies to human placenta paraffin sections. Histochem Cell Biol. 2008;130(3):595–599. doi: 10.1007/s00418-008-0458-z. PubMed DOI

Blaschitz A, Weiss U, Dohr G, Desoye G. Antibody reaction patterns in first trimester placenta: implications for trophoblast isolation and purity screening. Placenta. 2000;21(7):733–741. doi: 10.1053/plac.2000.0559. PubMed DOI

Bordeaux J, Welsh A, Agarwal S, et al. Antibody validation. Biotechniques. 2010;48(3):197–209. doi: 10.2144/000113382. PubMed DOI PMC

Bridges CC, Zalups RK. Mechanisms involved in the transport of mercuric ions in target tissues. Arch Toxicol. 2017;91(1):63–81. doi: 10.1007/s00204-016-1803-y. PubMed DOI PMC

Calatozzolo C, Gelati M, Ciusani E, et al. Expression of drug resistance proteins Pgp, MRP1, MRP3, MRP5 and GST-pi in human glioma. J Neurooncol. 2005;74(2):113–121. doi: 10.1007/s11060-004-6152-7. PubMed DOI

Cihalova D, Ceckova M, Kucera R, Klimes J, Staud F. Dinaciclib, a cyclin-dependent kinase inhibitor, is a substrate of human ABCB1 and ABCG2 and an inhibitor of human ABCC1 in vitro. Biochem Pharmacol. 2015;98(3):465–472. doi: 10.1016/j.bcp.2015.08.099. PubMed DOI

Clarkson TW. The three modern faces of mercury. Environ Health Perspect. 2002;110(Suppl 1):11–23. doi: 10.1289/ehp.02110s111. PubMed DOI PMC

Clarkson TW, Magos L. The toxicology of mercury and its chemical compounds. Crit Rev Toxicol. 2006;36(8):609–662. doi: 10.1080/10408440600845619. PubMed DOI

Cole SP. Multidrug resistance protein 1 (MRP1, ABCC1), a “multitasking” ATP-binding cassette (ABC) transporter. J Biol Chem. 2014;289(45):30880–30888. doi: 10.1074/jbc.R114.609248. PubMed DOI PMC

Cole SP, Bhardwaj G, Gerlach JH, et al. Overexpression of a transporter gene in a multidrug-resistant human lung cancer cell line. Science. 1992;258(5088):1650–1654. doi: 10.1126/science.1360704. PubMed DOI

Cole SP, Deeley RG. Transport of glutathione and glutathione conjugates by MRP1. Trends Pharmacol Sci. 2006;27(8):438–446. doi: 10.1016/j.tips.2006.06.008. PubMed DOI

Dallmann A, Liu XI, Burckart GJ, van den Anker J. Drug transporters expressed in the human placenta and models for studying maternal-fetal drug transfer. J Clin Pharmacol. 2019;59(Suppl 1):S70–S81. doi: 10.1002/jcph.1491. PubMed DOI PMC

Dartier J, Lemaitre E, Chourpa I, et al. ATP-dependent activity and mitochondrial localization of drug efflux pumps in doxorubicin-resistant breast cancer cells. Biochim Biophys Acta Gen Subj. 2017;1861(5 Pt A):1075–1084. doi: 10.1016/j.bbagen.2017.02.019. PubMed DOI

Ellison I, Richie JP., Jr Mechanisms of glutathione disulfide efflux from erythrocytes. Biochem Pharmacol. 2012;83(1):164–169. doi: 10.1016/j.bcp.2011.09.016. PubMed DOI

Evers R, Kool M, Smith AJ, van Deemter L, de Haas M, Borst P. Inhibitory effect of the reversal agents V-104, GF120918 and Pluronic L61 on MDR1 Pgp-, MRP1- and MRP2-mediated transport. Br J Cancer. 2000;83(3):366–374. doi: 10.1054/bjoc.2000.1260. PubMed DOI PMC

Evseenko D, Paxton JW, Keelan JA. Active transport across the human placenta: impact on drug efficacy and toxicity. Expert Opin Drug Met. 2006;2(1):51–69. doi: 10.1517/17425255.2.1.51. PubMed DOI

Evseenko DA, Paxton JW, Keelan JA. ABC drug transporter expression and functional activity in trophoblast-like cell lines and differentiating primary trophoblast. Am J Physiol Regul Integr Comp Physiol. 2006;290(5):R1357–R1365. doi: 10.1152/ajpregu.00630.2005. PubMed DOI

Farina M, Aschner M. Glutathione antioxidant system and methylmercury-induced neurotoxicity: An intriguing interplay. Biochim Biophys Acta Gen Subj. 2019;12:129285. doi: 10.1016/j.bbagen.2019.01.007. PubMed DOI PMC

Farina M, Aschner M, Rocha JB. Oxidative stress in MeHg-induced neurotoxicity. Toxicol Appl Pharmacol. 2011;256(3):405–417. doi: 10.1016/j.taap.2011.05.001. PubMed DOI PMC

Farina M, Avila DS, da Rocha JB, Aschner M. Metals, oxidative stress and neurodegeneration: a focus on iron, manganese and mercury. Neurochem Int. 2013;62(5):575–594. doi: 10.1016/j.neuint.2012.12.006. PubMed DOI PMC

Flens MJ, Izquierdo MA, Scheffer GL, et al. Immunochemical detection of the multidrug resistance-associated protein MRP in human multidrug-resistant tumor cells by monoclonal antibodies. Cancer Res. 1994;54(17):4557–4563. PubMed

Flens MJ, Zaman GJ, van der Valk P, et al. Tissue distribution of the multidrug resistance protein. Am J Pathol. 1996;148(4):1237–1247. PubMed PMC

Gennuso F, Fernetti C, Tirolo C, et al. Bilirubin protects astrocytes from its own toxicity by inducing up-regulation and translocation of multidrug resistance-associated protein 1 (Mrp1) Proc Natl Acad Sci U S A. 2004;101(8):2470–2475. doi: 10.1073/pnas.0308452100. PubMed DOI PMC

Giacomini KM, Huang SM, Tweedie DJ, et al. Membrane transporters in drug development. Nat Rev Drug Discov. 2010;9(3):215–236. doi: 10.1038/nrd3028. PubMed DOI PMC

Gundacker C, Hengstschlager M. The role of the placenta in fetal exposure to heavy metals. Wien Med Wochenschr. 2012;162(9–10):201–206. doi: 10.1007/s10354-012-0074-3. PubMed DOI

Gundacker C, Neesen J, Straka E, Ellinger I, Dolznig H, Hengstschläger M. Genetics of the human placenta: implications for toxicokinetics. Arch Toxicol. 2016;90(11):2563–2581. doi: 10.1007/s00204-016-1816-6. PubMed DOI

Gundacker C, Scheinast M, Damjanovic L, Fuchs C, Rosner M, Hengstschläger M. Proliferation potential of human amniotic fluid stem cells differently responds to mercury and lead exposure. Amino Acids. 2011;36(3):1–13. doi: 10.1007/s00726-011-1154-1. PubMed DOI

Hoffmeyer RE, Singh SP, Doonan CJ, et al. More on molecular mimicry in mercury toxicology. Chem Res Toxicol. 2006;19(9):1118–1120. doi: 10.1021/tx060177s. PubMed DOI

Jungsuwadee P, Nithipongvanitch R, Chen Y, et al. Mrp1 localization and function in cardiac mitochondria after doxorubicin. Mol Pharmacol. 2009;75(5):1117–1126. doi: 10.1124/mol.108.052209. PubMed DOI PMC

Kaufmann AM, Toro-Ramos AJ, Krise JP. Assessment of golgi apparatus versus plasma membrane-localized multi-drug resistance-associated protein 1. Mol Pharm. 2008;5(5):787–794. doi: 10.1021/mp8000205. PubMed DOI PMC

Kim HR, Lee KY, Ahn SG, et al. Transcriptional regulation, stabilization, and subcellular redistribution of multidrug resistance-associated protein 1 (MRP1) by glycogen synthase kinase 3alphabeta: novel insights on modes of cadmium-induced cell death stimulated by MRP1. Arch Toxicol. 2015;89(8):1271–1284. doi: 10.1007/s00204-014-1381-9. PubMed DOI

Kozlowska-Rup D, Czekaj P, Plewka D, Sikora J. Immunolocalization of ABC drug transporters in human placenta from normal and gestational diabetic pregnancies. Ginekol Pol. 2014;85(6):410–419. doi: 10.17772/gp/1745. PubMed DOI

Lang I, Pabst MA, Hiden U, et al. Heterogeneity of microvascular endothelial cells isolated from human term placenta and macrovascular umbilical vein endothelial cells. Eur J Cell Biol. 2003;82(4):163–173. doi: 10.1078/0171-9335-00306. PubMed DOI

Leong AS, Haffajee Z. Citraconic anhydride: a new antigen retrieval solution. Pathology. 2010;42(1):77–81. doi: 10.3109/00313020903434439. PubMed DOI

Long Y, Li Q, Cui Z. Molecular analysis and heavy metal detoxification of ABCC1/MRP1 in zebrafish. Mol Biol Rep. 2011;38(3):1703–1711. doi: 10.1007/s11033-010-0283-z. PubMed DOI

Maccani JZ, Koestler DC, Lester B, et al. Placental DNA methylation related to both infant toenail mercury and adverse neurobehavioral outcomes. Environ Health Perspect. 2015;123(7):723–729. doi: 10.1289/ehp.1408561. PubMed DOI PMC

Marchan R, Hammond CL, Ballatori N. Multidrug resistance-associated protein 1 as a major mediator of basal and apoptotic glutathione release. Biochim Biophys Acta. 2008;1778(10):2413–2420. doi: 10.1016/j.bbamem.2008.06.011. PubMed DOI PMC

Minich T, Riemer J, Schulz JB, Wielinga P, Wijnholds J, Dringen R. The multidrug resistance protein 1 (Mrp1), but not Mrp5, mediates export of glutathione and glutathione disulfide from brain astrocytes. J Neurochem. 2006;97(2):373–384. doi: 10.1111/j.1471-4159.2006.03737.x. PubMed DOI

Mueller CF, Widder JD, McNally JS, McCann L, Jones DP, Harrison DG. The role of the multidrug resistance protein-1 in modulation of endothelial cell oxidative stress. Circ Res. 2005;97(7):637–644. doi: 10.1161/01.RES.0000183734.21112.b7. PubMed DOI

Nagashige M, Ushigome F, Koyabu N, et al. Basal membrane localization of MRP1 in human placental trophoblast. Placenta. 2003;24(10):951–958. doi: 10.1016/S0143-4004(03)00170-X. PubMed DOI

Parry S, Zhang J. Multidrug resistance proteins affect drug transmission across the placenta. Am J Obstet Gynecol. 2007;196(5):476. doi: 10.1016/j.ajog.2007.02.019. PubMed DOI

Pascolo L, Fernetti C, Crivellato E, Mediavilla VMG, Ostrow DJ, Tiribelli C. MRP1 at apical (maternal-facing) pole of human trophoblasts accounts for the placental transfer of unconjugated bilirubin (UCB) from the fetal to the maternal circulation. J Hepatol. 2001;34:186–186. doi: 10.1016/S0168-8278(01)80685-1. DOI

Rajagopal A, Simon SM. Subcellular localization and activity of multidrug resistance proteins. Mol Biol Cell. 2003;14(8):3389–3399. doi: 10.1091/mbc.e02-11-0704. PubMed DOI PMC

Rosner M, Siegel N, Fuchs C, Slabina N, Dolznig H, Hengstschlager M. Efficient siRNA-mediated prolonged gene silencing in human amniotic fluid stem cells. Nat Protoc. 2010;5(6):1081–1095. doi: 10.1038/nprot.2010.74. PubMed DOI

Roundhill EA, Burchill SA. Detection and characterisation of multi-drug resistance protein 1 (MRP-1) in human mitochondria. Br J Cancer. 2012;106(6):1224–1233. doi: 10.1038/bjc.2012.40. PubMed DOI PMC

Rush T, Liu X, Nowakowski AB, Petering DH, Lobner D. Glutathione-mediated neuroprotection against methylmercury neurotoxicity in cortical culture is dependent on MRP1. Neurotoxicology. 2012;33(3):476–481. doi: 10.1016/j.neuro.2012.03.004. PubMed DOI

Sheehan MC, Burke TA, Navas-Acien A, Breysse PN, McGready J, Fox MA. Global methylmercury exposure from seafood consumption and risk of developmental neurotoxicity: a systematic review. Bull World Health Organ. 2014;92(4):254–269F. doi: 10.2471/BLT.12.116152. PubMed DOI PMC

Simmons-Willis TA, Koh AS, Clarkson TW, Ballatori N. Transport of a neurotoxicant by molecular mimicry: the methylmercury-L-cysteine complex is a substrate for human L-type large neutral amino acid transporter (LAT) 1 and LAT2. Biochem J. 2002;367(Pt 1):239–246. doi: 10.1042/BJ20020841. PubMed DOI PMC

St-Pierre MV, Serrano MA, Macias RI, et al. Expression of members of the multidrug resistance protein family in human term placenta. Am J Physiol Regul Integr Comp Physiol. 2000;279(4):R1495–R1503. doi: 10.1152/ajpregu.2000.279.4.R1495. PubMed DOI

Steinmeier J, Dringen R. Exposure of cultured astrocytes to menadione triggers rapid radical formation, glutathione oxidation and Mrp1-mediated export of glutathione disulfide. Neurochem Res. 2019;44(5):1167–1181. doi: 10.1007/s11064-019-02760-1. PubMed DOI

Straka E, Ellinger I, Balthasar C, et al. Mercury toxicokinetics of the healthy human term placenta involve amino acid transporters and ABC transporters. Toxicology. 2016;340:34–42. doi: 10.1016/j.tox.2015.12.005. PubMed DOI

Szentpetery Z, Sarkadi B, Bakos E, Varadi A. Functional studies on the MRP1 multidrug transporter: characterization of ABC-signature mutant variants. Anticancer Res. 2004;24(2A):449–455. PubMed

Tivnan A, Zakaria Z, O’Leary C, et al. Inhibition of multidrug resistance protein 1 (MRP1) improves chemotherapy drug response in primary and recurrent glioblastoma multiforme. Front Neurosci. 2015;9:218. doi: 10.3389/fnins.2015.00218. PubMed DOI PMC

Tucker EK, Nowak RA. Methylmercury alters proliferation, migration, and antioxidant capacity in human HTR8/SV-neo trophoblast cells. Reprod Toxicol. 2018;78:60–68. doi: 10.1016/j.reprotox.2018.03.008. PubMed DOI PMC

Van Luyn MJ, Muller M, Renes J, et al. Transport of glutathione conjugates into secretory vesicles is mediated by the multidrug-resistance protein 1. Int J Cancer. 1998;76(1):55–62. doi: 10.1002/(sici)1097-0215(19980330). PubMed DOI

Walker N, Filis P, Soffientini U, Bellingham M, O'Shaughnessy PJ, Fowler PA. Placental transporter localization and expression in the Human: the importance of species, sex, and gestational age differencesdagger. Biol Reprod. 2017;96(4):733–742. doi: 10.1093/biolre/iox012. PubMed DOI PMC

Wioland MA, Fleury-Feith J, Corlieu P, et al. CFTR, MDR1, and MRP1 immunolocalization in normal human nasal respiratory mucosa. J Histochem Cytochem. 2000;48(9):1215–1222. doi: 10.1177/002215540004800905. PubMed DOI

Yang L, Zhang Y, Wang F, Luo Z, Guo S, Strahle U. Toxicity of mercury: Molecular evidence. Chemosphere. 2020;245:125586. doi: 10.1016/j.chemosphere.2019.125586. PubMed DOI

Yin J, Zhang J. Multidrug resistance-associated protein 1 (MRP1/ABCC1) polymorphism: from discovery to clinical application. Zhong Nan Da Xue Xue Bao Yi Xue Ban. 2011;36(10):927–938. doi: 10.3969/j.issn.1672-7347.2011.10.002. PubMed DOI PMC

Zalups RK, Barfuss DW. Transport and toxicity of methylmercury along the proximal tubule of the rabbit. Toxicol Appl Pharmacol. 1993;121(2):176–185. doi: 10.1006/taap.1993.1143. PubMed DOI

Zhang Y, Han H, Elmquist WF, Miller DW. Expression of various multidrug resistance-associated protein (MRP) homologues in brain microvessel endothelial cells. Brain Res. 2000;876(1–2):148–153. doi: 10.1016/s0006-8993(00)02628-7. PubMed DOI

Zhao X, Li Y, Du K, et al. Involvement of human and canine MRP1 and MRP4 in benzylpenicillin transport. PLoS ONE. 2019;14(11):e0225702. doi: 10.1371/journal.pone.0225702. PubMed DOI PMC