Microscopy-based high-throughput assays enable multi-parametric analysis to assess adverse effects of nanomaterials in various cell lines
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Manufactured nanomaterials (MNMs) selected from a library of over 120 different MNMs with varied compositions, sizes, and surface coatings were tested by four different laboratories for toxicity by high-throughput/-content (HT/C) techniques. The selected particles comprise 14 MNMs composed of CeO2, Ag, TiO2, ZnO and SiO2 with different coatings and surface characteristics at varying concentrations. The MNMs were tested in different mammalian cell lines at concentrations between 0.5 and 250 µg/mL to link physical–chemical properties to multiple adverse effects. The cell lines are derived from relevant organs such as liver, lung, colon and the immune system. Endpoints such as viable cell count, cell membrane permeability, apoptotic cell death, mitochondrial membrane potential, lysosomal acidification and steatosis have been studied. Soluble MNMs, Ag and ZnO, were toxic in all cell types. TiO2 and SiO2 MNMs also triggered toxicity in some, but not all, cell types and the cell type-specific effects were influenced by the specific coating and surface modification. CeO2 MNMs were nearly ineffective in our test systems. Differentiated liver cells appear to be most sensitive to MNMs, Whereas most of the investigated MNMs showed no acute toxicity, it became clear that some show adverse effects dependent on the assay and cell line. Hence, it is advised that future nanosafety studies utilise a multi-parametric approach such as HT/C screening to avoid missing signs of toxicity. Furthermore, some of the cell type-specific effects should be followed up in more detail and might also provide an incentive to address potential adverse effects in vivo in the relevant organ.
KeywordsManufactured nanomaterials Toxicity High-throughput screening Cell type specificity Cell death Adverse outcome pathways Nanosafety
The authors acknowledge support from the European Commission’s 7th Framework Programme project NanoMILE (Contract No. NMP4-LA-2013-310451).
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
This article does not contain any studies with human participants or animals performed by any of the authors.
- Armand L, Tarantini A, Beal D, Biola-Clier M, Bobyk L, Sorieul S, Pernet-Gallay K, Marie-Desvergne C, Lynch I, Herlin-Boime N, Carriere M (2016) Long-term exposure of A549 cells to titanium dioxide nanoparticles induces DNA damage and sensitizes cells towards genotoxic agents. Nanotoxicology 10:913–923CrossRefPubMedGoogle Scholar
- Delaval M, Wohlleben W, Landsiedel R, Baeza-Squiban A, Boland S (2017) Assessment of the oxidative potential of nanoparticles by the cytochrome c assay: assay improvement and development of a high-throughput method to predict the toxicity of nanoparticles. Arch Toxicol 91:163–177CrossRefPubMedGoogle Scholar
- ECHA (2016) Proposal for harmonised classification and labelling. Substance name: Titanium dioxide. European Chemicals Agency. https://echa.europa.eu/documents/10162/594bf0e6-8789-4499-b9ba-59752f4eafab. Accessed 7 Nov 2017
- European commission (2013) Examination and assessment of consequences for industry, consumers, human health and the environment of possible options for changing the REACH requirements for nanomaterials. Final report. http://ec.europa.eu/environment/chemicals/nanotech/pdf/Final_Report.pdf. Joint Research Centre, Institute for Health and Consumer Protection, Reference: IHCP/2011/I/05/27/OC. Accessed 7 Nov 2017
- Godoy P, Hewitt NJ, Albrecht U, Andersen ME, Ansari N, Bhattacharya S, Bode JG, Bolleyn J, Borner C, Bottger J, Braeuning A, Budinsky RA, Burkhardt B, Cameron NR, Camussi G, Cho CS, Choi YJ, Craig RJ, Dahmen U, Damm G, Dirsch O, Donato MT, Dong J, Dooley S, Drasdo D, Eakins R, Ferreira KS, Fonsato V, Fraczek J, Gebhardt R, Gibson A, Glanemann M, Goldring CE, Gomez-Lechon MJ, Groothuis GM, Gustavsson L, Guyot C, Hallifax D, Hammad S, Hayward A, Haussinger D, Hellerbrand C, Hewitt P, Hoehme S, Holzhutter HG, Houston JB, Hrach J, Ito K, Jaeschke H, Keitel V, Kelm JM, Kevin PB, Kordes C, Kullak-Ublick GA, LeCluyse EL, Lu P, Luebke-Wheeler J, Lutz A, Maltman DJ, Matz-Soja M, McMullen P, Merfort I, Messner S, Meyer C, Mwinyi J, Naisbitt DJ, Nussler AK, Olinga P, Pampaloni F, Pi J, Pluta L, Przyborski SA, Ramachandran A, Rogiers V, Rowe C, Schelcher C, Schmich K, Schwarz M, Singh B, Stelzer EH, Stieger B, Stober R, Sugiyama Y, Tetta C, Thasler WE, Vanhaecke T, Vinken M, Weiss TS, Widera A, Woods CG, Xu JJ, Yarborough KM, Hengstler JG (2013) Recent advances in 2D and 3D in vitro systems using primary hepatocytes, alternative hepatocyte sources and non-parenchymal liver cells and their use in investigating mechanisms of hepatotoxicity, cell signaling and ADME. Arch Toxicol 87:1315–1530CrossRefPubMedPubMedCentralGoogle Scholar
- Hussain S, Thomassen LCJ, Ferecatu I, Borot MC, Andreau K, Martens JA, Fleury J, Baeza-Squiban A, Marano F, Boland S (2010) Carbon black and titanium dioxide nanoparticles elicit distinct apoptotic pathways in bronchial epithelial cells. Part Fibre Toxicol 7:10CrossRefPubMedPubMedCentralGoogle Scholar
- Jensen KA, Kembouche Y, Christiansen E, Jacobsen NR, Wallin H, Guiot C, Spalla O, Witschger O (2011) Final protocol for producing suitable manufactured nanomaterial exposure media—standard operation procedure (SPO) and background documentation. NanoGenoTox report. https://www.anses.fr/en/system/files/nanogenotox_deliverable_5.pdf. Accessed 7 Nov 2017
- Klein CL, Comero S, Stahlmecke B, Romazanov J, Kuhlbusch TAJ, Van Doren E, De Temmermann P-J, Mast J, Wick P, Krug H, Locoro G, Hund-Rinke K, Kördel W, Friedrichs S, Maier G, Werner J, Linsinger T, Gawlik BM (2011) NM-300 silver. Characterisation, stability, homogeneity. Publications Office of the European Union, European UnionGoogle Scholar
- Landesmann B (2015) Protein alkylation leading to liver fibrosis.aop wiki, https://aopwiki.org/wiki/images/c/c0/Aop38-Snapshot-March2015.pdf. Accessed 7 Nov 2017
- Landesmann B, Goumenou M, Munn S, Whelan M (2012) Description of prototype modes-of-action related to repeated dose toxicity. Publications Office of the European Union, European UnionGoogle Scholar
- Mennecozzi M, Landesmann B, Harris GA, Liska R, Whelan M (2012) Hepatotoxicity screening taking a mode-of-action approach using HepaRG cells and HCA. ALTEX Proc WC8 1:193–204Google Scholar
- Mülhopt S, Diabaté S, Krebs T, Weiss C, Paur HR (2009) Lung toxicity determination by in vitro exposure at the air-liquid interface with an integrated online dose measurement. J Phys 170:012008Google Scholar
- OECD (2010) Series on the safety of manufactured nanomaterials No. 27. List of manufactured nanomaterials and list of endpoints for phase one of the sponsorship programme for the testing of manufactured nanomaterials: revision. Organization for Economic Co-operation and Development (OECD), Environment directorate, ParisGoogle Scholar
- Ojea-Jimenez I, Urban P, Barahona F, Pedroni M, Capomaccio R, Ceccone G, Kinsner-Ovaskainen A, Rossi F, Gilliland D (2016) Highly flexible platform for tuning surface properties of silica nanoparticles and monitoring their biological interaction. ACS Appl Mater Interfaces 8:4838–4850CrossRefPubMedGoogle Scholar
- Panas A, Marquardt C, Nalcaci O, Bockhorn H, Baumann W, Paur HR, Mülhopt S, Diabaté S, Weiss C (2013) Screening of different metal oxide nanoparticles reveals selective toxicity and inflammatory potential of silica nanoparticles in lung epithelial cells and macrophages. Nanotoxicology 7:259–273CrossRefPubMedGoogle Scholar
- Panas A, Comouth A, Saathoff H, Leisner T, Al-Rawi M, Simon M, Seemann G, Dössel O, Mülhopt S, Paur HR, Fritsch-Decker S, Weiss C, Diabaté S (2014) Silica nanoparticles are less toxic to human lung cells when deposited at the air-liquid interface compared to conventional submerged exposure. Beilstein J Nanotechnol 5:1590–1602CrossRefPubMedPubMedCentralGoogle Scholar
- Piret JP, Bondarenko OM, Boyles MSP, Himly M, Ribeiro AR, Benetti F, Smal C, Lima B, Potthoff A, Simion M, Dumortier E, Leite PEC, Balottin LB, Granjeiro JM, Ivask A, Kahru A, Radauer-Preiml I, Tischler U, Duschl A, Saout C, Anguissola S, Haase A, Jacobs A, Nelissen I, Misra SK, Toussaint O (2017) Pan-European inter-laboratory studies on a panel of in vitro cytotoxicity and pro-inflammation assays for nanoparticles. Arch Toxicol 91:2315–2330CrossRefPubMedGoogle Scholar
- Rasmussen K, Mast J, De Temmerman PJ, Verleysen E, Waegeneers N, Van Steen F, Pizzolon JC, De Temmerman L, Van Doren E, Jensen KA, Birkedal R, Levin M, Nielsen SH, Koponen IK, Clausen PA, Kofoed-Sorensen V, Kembouche Y, Thieriet N, Spalla O, Guiot C, Rousset D, Witschger O, Bau S, Bianchi B, Motzkus C, Shivachev B, Dimova L, Nikolova R, Nihtianova D, Tarassov M, Petrov O, Bakardjieva S, Gilliland D, Pianella F, Ceccone G, Spampinato V, Cotogno G, Gibson N, Gaillard C, Mech A (2014) Titanium dioxide, NM-100, NM-101, NM-102, NM-103, NM-104, NM-105: characterisation and physico-chemical properties. Publications Office of the European Union, European UnionGoogle Scholar
- Sayes CM, Wahi R, Kurian PA, Liu Y, West JL, Ausman KD, Warheit DB, Colvin VL (2006) Correlating nanoscale titania structure with toxicity: a cytotoxicity and inflammatory response study with human dermal fibroblasts and human lung epithelial cells. Toxicol Sci 92:174–185CrossRefPubMedGoogle Scholar
- Singh C, Friedrichs S, Levin M, Birkedal R, Jensen KA, Pojana G, Wohlleben W, Schulte S, Wiench K, Turney T, Koulaeva O, Marshall D, Hund-Rinke K, Kördel W, Van Doren E, De Temmermann PJ, Abi Daoud Francisco M, Mast J, Gibson N, Koeber R, Linsinger T, Klein CL (2011) Zinc oxide NM-110, NM-111, NM-112, NM-113. Characterisation and test item preparation. Publications Office of the European Union, European UnionGoogle Scholar
- Singh C, Friedrichs S, Ceccone G, Gibson N, Jensen KA, Levin M, Infante HG, Carlander D, Rasmussen K (2014) Cerium dioxide, NM-211, NM-212, NM-213. Characterisation and test item preparation. Publications Office of the European Union, European UnionGoogle Scholar
- Tralau T, Oelgeschlager M, Gurtler R, Heinemeyer G, Herzler M, Hofer T, Itter H, Kuhl T, Lange N, Lorenz N, Muller-Graf C, Pabel U, Pirow R, Ritz V, Schafft H, Schneider H, Schulz T, Schumacher D, Zellmer S, Fleur-Bol G, Greiner M, Lahrssen-Wiederholt M, Lampen A, Luch A, Schonfelder G, Solecki R, Wittkowski R, Hensel A (2015) Regulatory toxicology in the twenty-first century: challenges, perspectives and possible solutions. Arch Toxicol 89:823–850CrossRefPubMedGoogle Scholar
- Yamashita K, Yoshioka Y, Higashisaka K, Mimura K, Morishita Y, Nozaki M, Yoshida T, Ogura T, Nabeshi H, Nagano K, Abe Y, Kamada H, Monobe Y, Imazawa T, Aoshima H, Shishido K, Kawai Y, Mayumi T, Tsunoda S, Itoh N, Yoshikawa T, Yanagihara I, Saito S, Tsutsumi Y (2011) Silica and titanium dioxide nanoparticles cause pregnancy complications in mice. Nat Nanotechnol 6:321–328CrossRefPubMedGoogle Scholar