Journal of Nanoparticle Research

, Volume 12, Issue 5, pp 1567–1578 | Cite as

Genomics-based screening of differentially expressed genes in the brains of mice exposed to silver nanoparticles via inhalation

  • Hye-Young Lee
  • You-Jin Choi
  • Eun-Jung Jung
  • Hu-Quan Yin
  • Jung-Taek Kwon
  • Ji-Eun Kim
  • Hwang-Tae Im
  • Myung-Haing Cho
  • Ju-Han Kim
  • Hyun-Young Kim
  • Byung-Hoon LeeEmail author
Research Paper


Silver nanoparticles (AgNP) are among the fastest growing product categories in the nanotechnology industry. Despite the importance of AgNP in consumer products and clinical applications, relatively little is known regarding AgNP toxicity and its associated risks. We investigated the effects of AgNP on gene expression in the mouse brain using Affymetrix Mouse Genome Arrays. C57BL/6 mice were exposed to AgNP (geometric mean diameter, 22.18 ± 1.72 nm; 1.91 × 107 particles/cm3) for 6 h/day, 5 days/week using the nose-only exposure system for 2 weeks. Total RNA isolated from the cerebrum and cerebellum was subjected to hybridization. From over 39,000 probe sets, 468 genes in the cerebrum and 952 genes in the cerebellum were identified as AgNP-responsive (one-way analysis of variance; p < 0.05). The largest groups of gene products affected by AgNP exposure included 73 genes in the cerebrum and 144 genes in the cerebellum. AgNP exposure modulated the expression of several genes associated with motor neuron disorders, neurodegenerative disease, and immune cell function, indicating potential neurotoxicity and immunotoxicity associated with AgNP exposure. Real-time PCR data for five genes analyzed from whole blood showed good correlation with the observed changes in the brain. Following rigorous validation and substantiation, these genes may assist in the development of surrogate markers for AgNP exposure and/or toxicity.


Brain Inhalation Mice Microarray Silver nanoparticles Toxicity EHS Nanomedicine 



This work was partially supported by the SRC/ERC program of MOST/KOSEF (R11-2007-107-01001-0).

Supplementary material

11051_2009_9666_MOESM1_ESM.jpg (275 kb)
Supplementary Data 1. Pathological analysis of the cerebrum obtained from C57BL/6 mice after inhalation exposure to AgNP. Histopathological analysis of hematoxylin-eosin-stained paraffin sections from (a) control and (b) AgNP cerebrum samples (×100). (JPG 275 kb)
11051_2009_9666_MOESM2_ESM.xls (91 kb)
Supplementary Data 2. Altered Genes compared with control (Bayesian ANOVA; p < 0.05) in the cerebrum. Values represent fold changes on log2 scale compared with control groups. (XLS 91 kb)
11051_2009_9666_MOESM3_ESM.xls (171 kb)
Supplementary Data 3. Altered Genes compared with control (Bayesian ANOVA; p < 0.05) in the cerebellum. Values represent fold changes on log2 scale compared with control groups. (XLS 171 kb)
11051_2009_9666_MOESM4_ESM.xls (28 kb)
Supplementary Data 4. Genes altered more than 1.5-fold (Bayesian ANOVA; p < 0.05) compared with control in the cerebrum. Values represent fold changes on log2 scale compared with control groups. (XLS 28 kb)
11051_2009_9666_MOESM5_ESM.xls (36 kb)
Supplementary Data 5. Genes altered more than 1.5-fold (Bayesian ANOVA; p < 0.05) compared with control in the cerebellum. Values represent fold changes on log2 scale compared with control groups. (XLS 37 kb)


  1. Aizawa H, Sekine Y, Takemura R, Zhang Z, Nangaku M, Hirokawa N (1992) Kinesin family in murine central nervous system. J Cell Biol 119:1287–1296. doi: 10.1083/jcb.119.5.1287 CrossRefPubMedGoogle Scholar
  2. Andreev SM, Babakhin AA, Petrukhina AO, Romanova VS, Parnes ZN, Petrov RV (2000) Immunogenic and allergenic properties of fullerene conjugates with amino acids and proteins. Dokl Biochem 370:4–7PubMedGoogle Scholar
  3. Block ML, Wu X, Pei Z, Li G, Wang T, Qin L, Wilson B, Yang J, Hong JS, Veronesi B (2004) Nanometer size diesel exhaust particles are selectively toxic to dopaminergic neurons: the role of microglia, phagocytosis, and NADPH oxidase. FASEB J 18:1618–1620PubMedGoogle Scholar
  4. Braydich-Stolle L, Hussain S, Schlager JJ, Hofmann MC (2005) In vitro cytotoxicity of nanoparticles in mammalian germline stem cells. Toxicol Sci 88:412–419. doi: 10.1093/toxsci/kfi256 CrossRefPubMedGoogle Scholar
  5. Cai H, Lin X, Xie C, Laird FM, Lai C, Wen H, Chiang HC, Shim H, Farah MH, Hoke A, Price DL, Wong PC (2005) Loss of ALS2 function is insufficient to trigger motor neuron degeneration in knock-out mice but predisposes neurons to oxidative stress. J Neurosci 25:7567–7574. doi: 10.1523/JNEUROSCI.1645-05.2005 CrossRefPubMedGoogle Scholar
  6. Cao YQ, Mantyh PW, Carlson EJ, Gillespie AM, Epstein CJ, Basbaum AI (1998) Primary afferent tachykinins are required to experience moderate to intense pain. Nature 392:390–394. doi: 10.1038/32897 CrossRefPubMedADSGoogle Scholar
  7. Chen X, Schluesener HJ (2008) Nanosilver: a nanoproduct in medical application. Toxicol Lett 176:1–12. doi: 10.1016/j.toxlet.2007.10.004 CrossRefPubMedGoogle Scholar
  8. Costa C, Tortosa R, Rodríguez A, Ferrer I, Torres JM, Bassols A, Pumarola M (2007) Aquaporin 1 and aquaporin 4 overexpression in bovine spongiform encephalopathy in a transgenic murine model and in cattle field cases. Brain Res 1175:96–106. doi: 10.1016/j.brainres.2007.06.088 CrossRefPubMedGoogle Scholar
  9. Czopik AK, Bynoe MS, Palm N, Raine CS, Medzhitov R (2006) Semaphorin 7A is a negative regulator of T cell responses. Immunity 24:591–600. doi: 10.1016/j.immuni.2006.03.013 CrossRefPubMedGoogle Scholar
  10. Dave KR, Raval AP, Purroy J, Kirkinezos IG, Moraes CT, Bradley WG, Pérez-Pinzón MA (2005) Aberrant delta PKC activation in the spinal cord of Wobbler mouse: a model of motor neuron disease. Neurobiol Dis 18:126–133. doi: 10.1016/j.nbd.2004.08.017 CrossRefPubMedGoogle Scholar
  11. Donaldson K, Brown D, Clouter A, Duffin R, MacNee W, Renwick L, Tran L, Stone V (2002) The pulmonary toxicology of ultrafine particles. J Aerosol Med 15:213–220. doi: 10.1089/089426802320282338 CrossRefPubMedGoogle Scholar
  12. Elder A, Gelein R, Silva V, Feikert T, Opanashuk L, Carter J, Potter R, Maynard A, Ito Y, Finkelstein J, Oberdörster G (2006) Translocation of inhaled ultrafine manganese oxide particles to the central nervous system. Environ Health Perspect 114:1172–1178CrossRefPubMedGoogle Scholar
  13. Feil R, Hartmann J, Luo C, Wolfsgruber W, Schilling K, Feil S, Barski JJ, Meyer M, Konnerth A, De Zeeuw CI, Hofmann F (2003) Impairment of LTD and cerebellar learning by Purkinje cell-specific ablation of cGMP-dependent protein kinase I. J Cell Biol 163:295–302. doi: 10.1083/jcb.200306148 CrossRefPubMedGoogle Scholar
  14. Fujita K, Morimoto Y, Ogami A, Myojyo T, Tanaka I, Shimada M, Wang WN, Endoh S, Uchida K, Nakazato T, Yamamoto K, Fukui H, Horie M, Yoshida Y, Iwahashi H, Nakanishi J (2009) Gene expression profiles in rat lung after inhalation exposure to C60 fullerene particles. Toxicology 258:47–55. doi: 10.1016/j.tox.2009.01.005 CrossRefPubMedGoogle Scholar
  15. Ghosh S, May MJ, Kopp EB (1998) NF-kappa B and Rel proteins: evolutionarily conserved mediators of immune responses. Annu Rev Immunol 16:225–260. doi: 10.1146/annurev.immunol.16.1.225 CrossRefPubMedGoogle Scholar
  16. Griffitt RJ, Hyndman K, Denslow ND, Barber DS (2009) Comparison of molecular and histological changes in zebrafish gills exposed to metallic nanoparticles. Toxicol Sci 107:404–415. doi: 10.1093/toxsci/kfn256 CrossRefPubMedGoogle Scholar
  17. Hoet PH, Brüske-Hohlfeld I, Salata OV (2004) Nanoparticles—known and unknown health risks. J Nanobiotechnol 2:12. doi: 10.1186/1477-3155-2-12 CrossRefGoogle Scholar
  18. Hussain SM, Hess KL, Gearhart JM, Geiss KT, Schlager JJ (2005) In vitro toxicity of nanoparticles in BRL 3A rat liver cells. Toxicol In Vitro 19:975–983. doi: 10.1016/j.tiv.2005.06.034 CrossRefPubMedGoogle Scholar
  19. Hussain SM, Javorina AK, Schrand AM, Duhart HM, Ali SF, Schlager JJ (2006) The interaction of manganese nanoparticles with PC-12 cells induces dopamine depletion. Toxicol Sci 92:456–463. doi: 10.1093/toxsci/kfl020 CrossRefPubMedGoogle Scholar
  20. International Council on Nanotechnology (2006) A Survey of Current Practices in the Nanotechnology Workplace. Available via DIALOG Accessed 31 October 2008
  21. Ji JH, Jung JH, Kim SS, Yoon JU, Park JD, Choi BS, Chung YH, Kwon IH, Jeong J, Han BS, Shin JH, Sung JH, Song KS, Yu IJ (2007) Twenty-eight-day inhalation toxicity study of silver nanoparticles in Sprague-Dawley rats. Inhal Toxicol 19:857–871. doi: 10.1080/08958370701432108 CrossRefPubMedGoogle Scholar
  22. Jiangxue W, Chunying C, Ying L, Fang J, Wei L, Fang L, Yufeng L, Bai L, Cuicui G, Guoqiang Z, Yuxi G, Yuliang Z, Zhifang C (2008a) Potential neurological lesion after nasal instillation of TiO2 nanoparticles in the anatase and rutile crystal phases. Toxicol Lett 183:72–80. doi: 10.1016/j.toxlet.2008.10.001 CrossRefGoogle Scholar
  23. Jiangxue W, Ying L, Fang J, Fang L, Wei L, Yiqun G, Yufeng L, Cuicui G, Guoqiang Z, Bai L, Yuliang Z, Zhifang C, Chunying C (2008b) Time-dependent translocation and potential impairment on central nervous system by intranasally instilled TiO2 nanoparticles. Toxicology 254:82–90. doi: 10.1016/j.tox.2008.09.014 CrossRefGoogle Scholar
  24. Jung JH, Oh HC, Noh HS, Ji JH, Kim SS (2006) Metal nanoparticle generation using a small ceramic heater with a local heating area. J Aerosol Sci 37:1662–1670. doi: 10.1016/j.jaerosci.2006.09.002 CrossRefGoogle Scholar
  25. Kanai Y, Okada Y, Tanaka Y, Harada A, Terada S, Hirokawa N (2000) KIF5C, a novel kinesin enriched in motor neurons. J Neurosci 20:6374–6384PubMedGoogle Scholar
  26. Kimura MY, Hosokawa H, Yamashita M, Hasegawa A, Iwamura C, Watarai H, Taniguchi M, Takagi T, Ishii S, Nakayama T (2005) Regulation of T helper type 2 cell differentiation by murine Schnurri-2. J Exp Med 201:397–408. doi: 10.1084/jem.20040733 CrossRefPubMedGoogle Scholar
  27. Kobayashi K, Takahashi M, Matsushita N, Miyazaki J, Koike M, Yaginuma H, Osumi N, Kaibuchi K, Kobayashi K (2004) Survival of developing motor neurons mediated by Rho GTPase signaling pathway through Rho-kinase. J Neurosci 24:3480–3488. doi: 10.1523/JNEUROSCI.0295-04.2004 CrossRefPubMedGoogle Scholar
  28. Kolodkin AL, Matthes DJ, Goodman CS (1993) The semaphorin genes encode a family of transmembrane and secreted growth cone guidance molecules. Cell 75:1389–1399. doi: 10.1016/0092-8674(93)90625-Z CrossRefPubMedGoogle Scholar
  29. Lansdown AB (2002) Silver. I: its antibacterial properties and mechanism of action. J Wound Care 11:125–130PubMedGoogle Scholar
  30. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) Method. Methods 25:402–408. doi: 10.1006/meth.2001.1262 CrossRefPubMedGoogle Scholar
  31. Long TC, Saleh N, Tilton RD, Lowry G, Veronesi B (2006) Titanium dioxide (P25) produces reactive oxygen species in immortalized brain microglia (BV2): implications for nanoparticle neurotoxicity. Environ Sci Technol 40:4346–4352. doi: 10.1021/es060589n CrossRefPubMedGoogle Scholar
  32. Morones JR, Elechiguerra JL, Camacho A, Kouri JB, Ramirez JT, Yacaman MJ (2005) The bactericidal effect of silver nanoparticles. Nanotechnology 16:2346–2353. doi: 10.1088/0957-4484/16/10/059 CrossRefADSGoogle Scholar
  33. Oberdörster G, Oberdörster E, Oberdörster J (2005) Nanotoxicology: an emerging discipline evolving from studies of ultrafine particles. Environ Health Perspect 113:823–839CrossRefPubMedGoogle Scholar
  34. Oukka M, Kim ST, Lugo G, Sun J, Wu LC, Glimcher LH (2002) A mammalian homolog of Drosophila schnurri, KRC, regulates TNF receptor-driven responses and interacts with TRAF2. Mol Cell 9:121–131. doi: 10.1016/S1097-2765(01)00434-8 CrossRefPubMedGoogle Scholar
  35. Peters A, Veronesi B, Calderón-Garcidueñas L, Gehr P, Chen LC, Geiser M, Reed W, Rothen-Rutishauser BM, Schürch S, Schulz H (2006) Translocation and potential neurological effects of fine and ultrafine particles a critical update. Part Fibre Toxicol 3:1–13. doi: 10.1186/1743-8977-3-13 CrossRefGoogle Scholar
  36. Roco MC (2005) Environmentally responsible development of nanotechnology. Environ Sci Technol 39:106A–112A. doi: 10.1021/es053199u CrossRefPubMedGoogle Scholar
  37. Rodríguez A, Pérez-Gracia E, Espinosa JC, Pumarola M, Torres JM, Ferrer I (2006) Increased expression of water channel aquaporin 1 and aquaporin 4 in Creutzfeldt-Jakob disease and in bovine spongiform encephalopathy-infected bovine-PrP transgenic mice. Acta Neuropathol 112:573–585. doi: 10.1007/s00401-006-0117-1 CrossRefPubMedGoogle Scholar
  38. Ruefli-Brasse AA, French DM, Dixit VM (2003) Regulation of NF-kappaB-dependent lymphocyte activation and development by paracaspase. Science 302:1581–1584. doi: 10.1126/science.1090769 CrossRefPubMedADSGoogle Scholar
  39. Stokin GB, Lillo C, Falzone TL, Brusch RG, Rockenstein E, Mount SL, Raman R, Davies P, Masliah E, Williams DS, Goldstein LS (2005) Axonopathy and transport deficits early in the pathogenesis of Alzheimer’s disease. Science 307:1282–1288. doi: 10.1126/science.1105681 CrossRefPubMedADSGoogle Scholar
  40. Takagi T, Harada J, Ishii S (2001) Murine Schnurri-2 is required for positive selection of thymocytes. Nat Immunol 2:1048–1053. doi: 10.1038/ni728 CrossRefPubMedGoogle Scholar
  41. Tin-Tin-Win-Shwe, Mitsushima D, Yamamoto S, Fukushima A, Funabashi T, Kobayashi T, Fujimaki H (2008) Changes in neurotransmitter levels and proinflammatory cytokine mRNA expression in the mice olfactory bulb following nanoparticle exposure. Toxicol Appl Pharmacol 226:192–198. doi: 10.1016/j.taap.2007.09.009 CrossRefPubMedGoogle Scholar
  42. Woodrow Wilson International Center for Scholars (2007) Nanotechnology Consumer Products Inventory. Available via DIALOG Accessed 31 October 2008
  43. Yamanaka K, Vande Velde C, Eymard-Pierre E, Bertini E, Boespflug-Tanguy O, Cleveland DW (2003) Unstable mutants in the peripheral endosomal membrane component ALS2 cause early-onset motor neuron disease. Proc Natl Acad Sci USA 100:16041–16046. doi: 10.1073/pnas.2635267100 CrossRefPubMedADSGoogle Scholar
  44. Yamanaka K, Miller TM, McAlonis-Downes M, Chun SJ, Cleveland DW (2006) Progressive spinal axonal degeneration and slowness in ALS2-deficient mice. Ann Neurol 60:95–104. doi: 10.1002/ana.20888 CrossRefPubMedGoogle Scholar
  45. Zhang T, Stilwell JL, Gerion D, Ding L, Elboudwarej O, Cooke PA, Gray JW, Alivisatos AP, Chen FF (2006) Cellular effect of high doses of silica-coated quantum dot profiled with high throughput gene expression analysis and high content cellomics measurements. Nano Lett 6:800–808. doi: 10.1021/nl0603350 CrossRefPubMedADSGoogle Scholar
  46. Zimmer A, Zimmer AM, Baffi J, Usdin T, Reynolds K, König M, Palkovits M, Mezey E (1998) Hypoalgesia in mice with a targeted deletion of the tachykinin 1 gene. Proc Natl Acad Sci USA 95:2630–2635. doi: 10.1073/pnas.95.5.2630 CrossRefPubMedADSGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2009

Authors and Affiliations

  • Hye-Young Lee
    • 1
  • You-Jin Choi
    • 1
  • Eun-Jung Jung
    • 1
  • Hu-Quan Yin
    • 1
  • Jung-Taek Kwon
    • 2
  • Ji-Eun Kim
    • 2
    • 4
  • Hwang-Tae Im
    • 2
    • 4
  • Myung-Haing Cho
    • 2
    • 4
  • Ju-Han Kim
    • 3
  • Hyun-Young Kim
    • 5
  • Byung-Hoon Lee
    • 1
    Email author
  1. 1.College of Pharmacy and Research Institute of Pharmaceutical SciencesSeoul National UniversitySeoulRepublic of Korea
  2. 2.College of Veterinary MedicineSeoul National UniversitySeoulRepublic of Korea
  3. 3.College of MedicineSeoul National UniversitySeoulRepublic of Korea
  4. 4.Nano Systems Institute-National Core Research CenterSeoul National UniversitySeoulRepublic of Korea
  5. 5.Chemical Safety and Health Research CenterOccupational Safety and Health Research InstituteDaejeonRepublic of Korea

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