Abstract
The extensive applications of nanoparticles due to their unique physicochemical and biological properties have resulted in a surge in production and usage. The large-scale production and application of nanomaterials can result in their exposure to various living beings. The nanoparticle exposure can be intentional or unintentional. Many nanoparticles have a toxic response due to their increased bioactivity compared to the bulk counterpart. Hence, it is necessary to evaluate the toxicity associated with the nanoparticle. A wide spectrum of toxicity assessment tests is available and categorized into in vitro and in vivo assays. In vitro toxicity assays measure the cell’s metabolic function, membrane integrity, protein content, organelles function, and various enzymatic activities. Alternatively, in vivo toxicity assays evaluate organ-level toxicity and toxicokinetics. The selection of cell lines to perform in vitro toxicity and animal models for in vivo toxicity is critical. This chapter summarizes the various toxicity assays performed to evaluate the toxicity associated with the nanoparticle treatments.
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References
Ahamed M, Posgai R, Gorey TJ, Nielsen M, Hussain SM, Rowe JJ (2010) Silver nanoparticles induced heat shock protein 70, oxidative stress and apoptosis in Drosophila melanogaster. Toxicol Appl Pharmacol 242:263–269. https://doi.org/10.1016/j.taap.2009.10.016
Ahmed B, Dwivedi S, Abdin MZ, Azam A, Al-Shaeri M, Khan MS, Saquib Q, Al-Khedhairy AA, Musarrat J (2017) Mitochondrial and chromosomal damage induced by oxidative stress in Zn(2+) ions, ZnO-bulk and ZnO-NPs treated allium cepa roots. Sci Rep 7:40685. https://doi.org/10.1038/srep40685
Alaraby M, Hernández A, Marcos R (2017) Copper oxide nanoparticles and copper sulphate act as antigenotoxic agents in drosophila melanogaster. Environ Mol Mutagen 58:46–55. https://doi.org/10.1002/em.22068
Andreotti PE, Cree IA, Kurbacher CM, Hartmann DM, Linder D, Harel G, Gleiberman I, Caruso PA, Ricks SH, Untch M (1995) Chemosensitivity testing of human tumors using a microplate adenosine triphosphate luminescence assay: clinical correlation for cisplatin resistance of ovarian carcinoma. Cancer Res 55:5276–5282
Anoopkumar-Dukie S, Carey JB, Conere T, O’Sullivan E, van Pelt FN, Allshire A (2005) Resazurin assay of radiation response in cultured cells. Br J Radiol 78:945–947. https://doi.org/10.1259/bjr/54004230
AshaRani PV, Mun GLK, Hande MP, Valiyaveettil S (2009) Cytotoxicity and genotoxicity of silver nanoparticles in human cells. ACS Nano 3:279–290. https://doi.org/10.1021/nn800596w
Aspatwar A, Hammaren MM, Parikka M, Parkkila S (2019) Rapid evaluation of toxicity of chemical compounds using zebrafish embryos. J Vis Exp. https://doi.org/10.3791/59315
Ates G, Vanhaecke T, Rogiers V, Rodrigues RM (2017) Assaying cellular viability using the neutral red uptake assay. Methods Mol Biol 1601:19–26. https://doi.org/10.1007/978-1-4939-6960-9_2
Avlasevich S, Bryce S, De Boeck M, Elhajouji A, Van Goethem F, Lynch A, Nicolette J, Shi J, Dertinger S (2011) Flow cytometric analysis of micronuclei in mammalian cell cultures: past, present and future. Mutagenesis 26:147–152. https://doi.org/10.1093/mutage/geq058
Barreto A, Carvalho A, Campos A, Osório H, Pinto E, Almeida A, Trindade T, Soares A, Hylland K, Loureiro S (2020) Effects of gold nanoparticles in gilthead seabream—a proteomic approach. Aquat Toxicol 221:105445
Boran H, Ulutas G (2016) Genotoxic effects and gene expression changes in larval zebrafish after exposure to ZnCl2 and ZnO nanoparticles. Dis Aquat Org 117:205–214. https://doi.org/10.3354/dao02943
Bowman L, Castranova V, Ding M (2012) Single cell gel electrophoresis assay (comet assay) for evaluating nanoparticles-induced DNA damage in cells. Methods Mol Biol 906:415–422. https://doi.org/10.1007/978-1-61779-953-2_34
Bratosin D, Mitrofan L, Palii C, Estaquier J, Montreuil J (2005) Novel fluorescence assay using calcein-AM for the determination of human erythrocyte viability and aging. Cytometry A 66A:78–84. https://doi.org/10.1002/cyto.a.20152
Braun K, Stürzel CM, Biskupek J, Kaiser U, Kirchhoff F, Lindén M (2018) Comparison of different cytotoxicity assays for in vitro evaluation of mesoporous silica nanoparticles. Toxicol In Vitro 52:214–221. https://doi.org/10.1016/j.tiv.2018.06.019
Braydich-Stolle L, Hussain S, Schlager JJ, Hofmann M-C (2005) In vitro cytotoxicity of nanoparticles in mammalian germline stem cells. Toxicol Sci 88:412–419. https://doi.org/10.1093/toxsci/kfi256
Buehler EV (1965) Delayed contact hypersensitivity in the guinea pig. Arch Dermatol 91:171–177. https://doi.org/10.1001/archderm.1965.01600080079017
Carmona ER, Inostroza-Blancheteau C, Rubio L, Marcos R (2016) Genotoxic and oxidative stress potential of nanosized and bulk zinc oxide particles in Drosophila melanogaster. Toxicol Ind Health 32:1987–2001. https://doi.org/10.1177/0748233715599472
Chen Y-S, Hung Y-C, Liau I, Huang GS (2009) Assessment of the in vivo toxicity of gold nanoparticles. Nanoscale Res Lett 4:858. https://doi.org/10.1007/s11671-009-9334-6
Correia B, Lourenco J, Marques S, Nogueira V, Gavina A, da Graça Rasteiro M, Antunes F, Mendo S, Pereira R (2017) Oxidative stress and genotoxicity of an organic and an inorganic nanomaterial to Eisenia andrei: SDS/DDAB nano-vesicles and titanium silicon oxide. Ecotoxicol Environ Saf 140:198–205
Correia AT, Rodrigues S, Ferreira-Martins D, Nunes AC, Ribeiro MI, Antunes SC (2020) Multi-biomarker approach to assess the acute effects of cerium dioxide nanoparticles in gills, liver and kidney of Oncorhynchus mykiss. Comp Biochem Physiol C Toxicol Pharmacol 238:108842
Czekanska EM (2011) Assessment of cell proliferation with resazurin-based fluorescent dye. Methods Mol Biol 740:27–32. https://doi.org/10.1007/978-1-61779-108-6_5
de Alteriis E, Falanga A, Galdiero S, Guida M, Maselli V, Galdiero E (2018) Genotoxicity of gold nanoparticles functionalised with indolicidin towards Saccharomyces cerevisiae. J Environ Sci 66:138–145. https://doi.org/10.1016/j.jes.2017.04.034
de Souza Trigueiro NS, Gonçalves BB, Dias FC, de Oliveira Lima EC, Rocha TL, Sabóia-Morais SMT (2021) Co-exposure of iron oxide nanoparticles and glyphosate-based herbicide induces DNA damage and mutagenic effects in the guppy (Poecilia reticulata). Environ Toxicol Pharmacol 81:103521. https://doi.org/10.1016/j.etap.2020.103521
Demir E (2020) An in vivo study of nanorod, nanosphere, and nanowire forms of titanium dioxide using Drosophila melanogaster: toxicity, cellular uptake, oxidative stress, and DNA damage. J Toxicol Environ Health A 83:456–469. https://doi.org/10.1080/15287394.2020.1777236
Eckrich J, Kugler P, Buhr CR, Ernst BP, Mendler S, Baumgart J, Brieger J, Wiesmann N (2020) Monitoring of tumor growth and vascularisation with repetitive ultrasonography in the chicken chorioallantoic-membrane-assay. Sci Rep 10:1–14
Fenech M (2008) The micronucleus assay determination of chromosomal level DNA damage. Methods Mol Biol 410:185–216. https://doi.org/10.1007/978-1-59745-548-0_12
Frankild S, Vølund A, Wahlberg JE, Andersen KE (2000) Comparison of the sensitivities of the Buehler test and the guinea pig maximisation test for predictive testing of contact allergy. Acta Derm Venereol 80:256–262. https://doi.org/10.1080/000155500750012126
George JM, Magogotya M, Vetten MA, Buys AV, Gulumian M (2017) From the cover: an investigation of the genotoxicity and interference of gold nanoparticles in commonly used in vitro mutagenicity and genotoxicity assays. Toxicol Sci 156:149–166. https://doi.org/10.1093/toxsci/kfw247
Gerberick GF, Ryan CA, Dearman RJ, Kimber I (2007) Local lymph node assay (LLNA) for detection of sensitisation capacity of chemicals. Methods 41:54–60. https://doi.org/10.1016/j.ymeth.2006.07.006
Ghosh I, Sadhu A, Moriyasu Y, Bandyopadhyay M, Mukherjee A (2019) Manganese oxide nanoparticles induce genotoxicity and DNA hypomethylation in the moss Physcomitrella patens. Mutat Res 842:146–157. https://doi.org/10.1016/j.mrgentox.2018.12.006
Grodzik M, Sawosz E (2006) The influence of silver nanoparticles on chicken embryo development and bursa of Fabricius morphology. J Anim Feed Sci 15:111–114. https://doi.org/10.22358/jafs/70155/2006
Hansen MB, Nielsen SE, Berg K (1989) Re-examination and further development of a precise and rapid dye method for measuring cell growth/cell kill. J Immunol Methods 119:203–210
Hayashi M (2016) The micronucleus test—most widely used in vivo genotoxicity test. Genes Environ 38:18. https://doi.org/10.1186/s41021-016-0044-x
Hu Y, Chen X, Yang K, Lin D (2018) Distinct toxicity of silver nanoparticles and silver nitrate to Daphnia magna in M4 medium and surface water. Sci Total Environ 618:838–846. https://doi.org/10.1016/j.scitotenv.2017.08.222
Huang KT, Chen YH, Walker AM (2004) Inaccuracies in MTS assays: major distorting effects of medium, serum albumin, and fatty acids. BioTechniques 37:406–412. https://doi.org/10.2144/04373ST05
Huyck L, Ampe C, Van Troys M (2012) The XTT cell proliferation assay applied to cell layers embedded in three-dimensional matrix. Assay Drug Dev Technol 10:382–392. https://doi.org/10.1089/adt.2011.391
Ishiyama M, Tominaga H, Shiga M, Sasamoto K, Ohkura Y, Ueno K (1996) A combined assay of cell vability and in vitro cytotoxicity with a highly water-soluble tetrazolium salt, neutral red and crystal violet. Biol Pharm Bull 19:1518–1520
Jia Y-P, Ma B-Y, Wei X-W, Qian Z-Y (2017) The in vitro and in vivo toxicity of gold nanoparticles. Chin Chem Lett 28:691–702. https://doi.org/10.1016/j.cclet.2017.01.021
Jonsson B, Liminga G, Csoka K, Fridborg H, Dhar S, Nygren P, Larsson R (1996) Cytotoxic activity of calcein acetoxymethyl ester (Calcein/AM) on primary cultures of human haematological and solid tumours. Eur J Cancer 32:883–887
Joseph X, Akhil V, Arathi A, Mohanan PV (2021) Comprehensive development in organ-on-a-chip technology. J Pharm Sci. https://doi.org/10.1016/j.xphs.2021.07.014
Kaja S, Payne AJ, Naumchuk Y, Koulen P (2017) Quantification of lactate dehydrogenase for cell viability testing using cell lines and primary cultured astrocytes. Curr Protoc Toxicol 72:2.26.1–2.26.10. https://doi.org/10.1002/cptx.21
Keepers YP, Pizao PE, Peters GJ, van Ark-Otte J, Winograd B, Pinedo HM (1991) Comparison of the sulforhodamine B protein and tetrazolium (MTT) assays for in vitro chemosensitivity testing. Eur J Cancer Clin Oncol 27:897–900. https://doi.org/10.1016/0277-5379(91)90142-Z
Kendig DM, Tarloff JB (2007) Inactivation of lactate dehydrogenase by several chemicals: implications for in vitro toxicology studies. Toxicol In Vitro 21:125–132. https://doi.org/10.1016/j.tiv.2006.08.004
Kimmel CB, Ballard WW, Kimmel SR, Ullmann B, Schilling TF (1995) Stages of embryonic development of the zebrafish. Dev Dyn 203:253–310. https://doi.org/10.1002/aja.1002030302
Koehlé-Divo V, Cossu-Leguille C, Pain-Devin S, Simonin C, Bertrand C, Sohm B, Mouneyrac C, Devin S, Giambérini L (2018) Genotoxicity and physiological effects of CeO2 NPs on a freshwater bivalve (Corbicula fluminea). Aquat Toxicol 198:141–148
Krishnaraj C, Harper SL, Yun S-I (2016) In vivo toxicological assessment of biologically synthesised silver nanoparticles in adult Zebrafish (Danio rerio). J Hazard Mater 301:480–491. https://doi.org/10.1016/j.jhazmat.2015.09.022
Kumar V, Sharma N, Maitra SS (2017) In vitro and in vivo toxicity assessment of nanoparticles. Int Nano Lett 7:243–256
Kumar P, Nagarajan A, Uchil PD (2018) Analysis of cell viability by the lactate dehydrogenase assay. Cold Spring Harb Protoc 2018. https://doi.org/10.1101/pdb.prot095497
Kurbacher CM, Cree IA (2005) Chemosensitivity testing using microplate adenosine triphosphate-based luminescence measurements. Methods Mol Med 110:101–120. https://doi.org/10.1385/1-59259-869-2:101
Lei R, Wu C, Yang B, Ma H, Shi C, Wang Q, Wang Q, Yuan Y, Liao M (2008) Integrated metabolomic analysis of the nano-sized copper particle-induced hepatotoxicity and nephrotoxicity in rats: a rapid in vivo screening method for nanotoxicity. Toxicol Appl Pharmacol 232:292–301
Liminga G, Nygren P, Dhar S, Nilsson K, Larsson R (1995) Cytotoxic effect of calcein acetoxymethyl ester on human tumor cell lines: drug delivery by intracellular trapping. Anticancer Drugs 6:578–585
Lorenzo Y, Costa S, Collins AR, Azqueta A (2013) The comet assay, DNA damage, DNA repair and cytotoxicity: hedgehogs are not always dead. Mutagenesis 28:427–432. https://doi.org/10.1093/mutage/get018
Luepke NP (1985) Hen’s egg chorioallantoic membrane test for irritation potential. Food Chem Toxicol 23:287–291. https://doi.org/10.1016/0278-6915(85)90030-4
Maehara Y, Anai H, Tamada R, Sugimachi K (1987) The ATP assay is more sensitive than the succinate dehydrogenase inhibition test for predicting cell viability. Eur J Cancer Clin Oncol 23:273–276. https://doi.org/10.1016/0277-5379(87)90070-8
Magnusson B (1980) Identification of contact sensitisers by animal assay. Contact Dermatitis 6:46–50. https://doi.org/10.1111/j.1600-0536.1980.tb03894.x
Magro M, De Liguoro M, Franzago E, Baratella D, Vianello F (2018) The surface reactivity of iron oxide nanoparticles as a potential hazard for aquatic environments: a study on Daphnia magna adults and embryos. Sci Rep 8:13017. https://doi.org/10.1038/s41598-018-31483-6
Mahjoubian M, Naeemi AS, Sheykhan M (2021) Toxicological effects of Ag(2)O and Ag(2)CO(3) doped TiO(2) nanoparticles and pure TiO(2) particles on zebrafish (Danio rerio). Chemosphere 263:128182. https://doi.org/10.1016/j.chemosphere.2020.128182
Mosmann T (1983) Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods 65:55–63
Murugadas A, Zeeshan M, Thamaraiselvi K, Ghaskadbi S, Akbarsha MA (2016) Hydra as a model organism to decipher the toxic effects of copper oxide nanorod: eco-toxicogenomics approach. Sci Rep 6:29663. https://doi.org/10.1038/srep29663
Ngamwongsatit P, Banada PP, Panbangred W, Bhunia AK (2008) WST-1-based cell cytotoxicity assay as a substitute for MTT-based assay for rapid detection of toxigenic Bacillus species using CHO cell line. J Microbiol Methods 73:211–215. https://doi.org/10.1016/j.mimet.2008.03.002
OECD (1994) OECD guidelines for the testing of chemicals. Organization for Economic, Paris
OECD (2016a) Test No. 487: In vitro mammalian cell micronucleus test. https://doi.org/10.1787/9789264264861-en
OECD (2016b) Test No. 474: Mammalian erythrocyte micronucleus test. https://doi.org/10.1787/9789264264762-en
OECD (2016c) Test No. 473: In vitro mammalian chromosomal aberration test. https://doi.org/10.1787/9789264264649-en
OECD (2016d) Test No. 475: Mammalian bone marrow chromosomal aberration test. https://doi.org/10.1787/9789264264786-en
Oliviero M, Schiavo S, Dumontet S, Manzo S (2019) DNA damages and offspring quality in sea urchin Paracentrotus lividus sperms exposed to ZnO nanoparticles. Sci Total Environ 651:756–765. https://doi.org/10.1016/j.scitotenv.2018.09.243
Patel S, Patel P, Bakshi SR (2017) Titanium dioxide nanoparticles: an in vitro study of DNA binding, chromosome aberration assay, and comet assay. Cytotechnology 69:245–263. https://doi.org/10.1007/s10616-016-0054-3
Pompa PP, Vecchio G, Galeone A, Brunetti V, Sabella S, Maiorano G, Falqui A, Bertoni G, Cingolani R (2011) In vivo toxicity assessment of gold nanoparticles in Drosophila melanogaster. Nano Res 4:405–413
Prasek M, Sawosz E, Jaworski S, Grodzik M, Ostaszewska T, Kamaszewski M, Wierzbicki M, Chwalibog A (2013) Influence of nanoparticles of platinum on chicken embryo development and brain morphology. Nanoscale Res Lett 8:251. https://doi.org/10.1186/1556-276X-8-251
Ramirez CN, Antczak C, Djaballah H (2010) Cell viability assessment: toward content-rich platforms. Expert Opin Drug Discov 5:223–233. https://doi.org/10.1517/17460441003596685
Repetto G, del Paso A, Zurita JL (2008) Neutral red uptake assay for the estimation of cell viability/cytotoxicity. Nat Protoc 3:1125–1131
Ribatti D (2016) The chick embryo chorioallantoic membrane (CAM). A multifaceted experimental model. Mech Dev 141:70–77
Roehm NW, Rodgers GH, Hatfield SM, Glasebrook AL (1991) An improved colorimetric assay for cell proliferation and viability utilising the tetrazolium salt XTT. J Immunol Methods 142:257–265
Roman D, Yasmeen A, Mireuta M, Stiharu I, Al Moustafa A-E (2013) Significant toxic role for single-walled carbon nanotubes during normal embryogenesis. Nanomedicine 9:945–950. https://doi.org/10.1016/j.nano.2013.03.010
Rovida C, Ryan C, Cinelli S, Basketter D, Dearman R, Kimber I (2012) The local lymph node assay (LLNA). Curr Protoc Toxicol Chapter 20:Unit 20.7. https://doi.org/10.1002/0471140856.tx2007s51
Rubinstein LV, Shoemaker RH, Paull KD, Simon RM, Tosini S, Skehan P, Scudiero DA, Monks A, Boyd MR (1990) Comparison of in vitro anticancer-drug-screening data generated with a tetrazolium assay versus a protein assay against a diverse panel of human tumor cell lines. JNCI J Natl Cancer Inst 82:1113–1117. https://doi.org/10.1093/jnci/82.13.1113
Savić-Zdravković D, Milošević D, Uluer E, Duran H, Matić S, Stanić S, Vidmar J, Ščančar J, Dikic D, Jovanović B (2020) A multiparametric approach to cerium oxide nanoparticle toxicity assessment in non-biting midges. Environ Toxicol Chem 39:131–140. https://doi.org/10.1002/etc.4605
Singh Z, Singh I (2019) CTAB surfactant assisted and high pH nano-formulations of CuO nanoparticles pose greater cytotoxic and genotoxic effects. Sci Rep 9:5880. https://doi.org/10.1038/s41598-019-42419-z
Skehan P, Storeng R, Scudiero D, Monks A, McMahon J, Vistica D, Warren JT, Bokesch H, Kenney S, Boyd MR (1990) New colorimetric cytotoxicity assay for anticancer-drug screening. JNCI J Natl Cancer Inst 82:1107–1112. https://doi.org/10.1093/jnci/82.13.1107
Suriyaprabha R, Balu KS, Karthik S, Prabhu M, Rajendran V, Aicher WK, Maaza M (2019) A sensitive refining of in vitro and in vivo toxicological behavior of green synthesised ZnO nanoparticles from the shells of Jatropha curcas for multifunctional biomaterials development. Ecotoxicol Environ Saf 184:109621. https://doi.org/10.1016/j.ecoenv.2019.109621
Thomas P, Fenech M (2011) Cytokinesis-block micronucleus cytome assay in lymphocytes. Methods Mol Biol 682:217–234. https://doi.org/10.1007/978-1-60327-409-8_16
Truong L, Harper SL, Tanguay RL (2011) Evaluation of embryotoxicity using the zebrafish model. Methods Mol Biol 691:271–279. https://doi.org/10.1007/978-1-60761-849-2_16
Van Rensburg CE, Anderson R, Jooné G, Myer MS, O’Sullivan JF (1997) Novel tetramethylpiperidine-substituted phenazines are potent inhibitors of P-glycoprotein activity in a multidrug resistant cancer cell line. Anticancer Drugs 8:708–713
Vandghanooni S, Eskandani M (2011) Comet assay: a method to evaluate genotoxicity of nano-drug delivery system. Bioimpacts 1:87–97. https://doi.org/10.5681/bi.2011.012
Villacis RAR, José Filho S, Pina B, Azevedo RB, Pic-Taylor A, Mazzeu JF, Grisolia CK (2017) Integrated assessment of toxic effects of maghemite (γ-Fe2O3) nanoparticles in zebrafish. Aquat Toxicol 191:219–225
Vu BT, Shahin SA, Croissant J, Fatieiev Y, Matsumoto K, Le-Hoang Doan T, Yik T, Simargi S, Conteras A, Ratliff L, Jimenez CM, Raehm L, Khashab N, Durand J-O, Glackin C, Tamanoi F (2018) Chick chorioallantoic membrane assay as an in vivo model to study the effect of nanoparticle-based anticancer drugs in ovarian cancer. Sci Rep 8:8524. https://doi.org/10.1038/s41598-018-25573-8
Xu EG, Cheong RS, Liu L, Hernandez LM, Azimzada A, Bayen S, Tufenkji N (2020) Primary and secondary plastic particles exhibit limited acute toxicity but chronic effects on Daphnia magna. Environ Sci Technol 54:6859–6868. https://doi.org/10.1021/acs.est.0c00245
Zhao C-M, Wang W-X (2011) Comparison of acute and chronic toxicity of silver nanoparticles and silver nitrate to Daphnia magna. Environ Toxicol Chem 30:885–892. https://doi.org/10.1002/etc.451
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The authors wish to express their sincere thanks to the Director and Head, Biomedical Technology Wing, Sree Chitra Tirunal Institute for Medical Sciences and Technology (Govt. of India), Trivandrum, for their support and for providing the infrastructure to carry out this work.
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Joseph, X., Akhil, Arathi, Megha, K.B., Vandana, U., Mohanan, P.V. (2023). Toxicity Assessment of Nanoparticle. In: Mohanan, P.V., Kappalli, S. (eds) Biomedical Applications and Toxicity of Nanomaterials. Springer, Singapore. https://doi.org/10.1007/978-981-19-7834-0_16
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