title: Investigating the immunomodulatory nature of zinc oxide nanoparticles at sub-cytotoxic levels in vitro and after intranasal instillation in vivo.
abstract: BACKGROUND
  This study evaluates the time-dependent pro-inflammatory response of the model human lung epithelial cells (A549) to industrially relevant zinc oxide nanoparticles (ZnO NPs). In terms of toxicity, ZnO-NPs are categorised into the group of high toxicity nanomaterials. However information on pro-inflammatory potential of these NPs at sub-toxic concentrations is limited. Understanding how cellular defense mechanisms function in the presence of sub-cytotoxic concentrations of these NPs is vital. Moreover, there is an urgent need for additional in vivo studies addressing pulmonary toxicity due to accidental inhalation of ZnO NPs.
  
  
  RESULTS
  Exposure to sub-cytotoxic ZnO NP concentrations (20 μg/mL) induced significant up-regulation of mRNA for the pro-inflammatory cytokine IL-8 and redox stress marker heme oxygenase-1, along with increased release of IL-8. The highest pro-inflammatory response was recorded after 4 to 6 hr exposure to ZnO NPs over a 24 hr period. Pre-treatment of A549 cells with the sulfhydryl antioxidant N-acetyl cysteine (at 5 mM) resulted in significant reduction of the up-regulation of inflammatory markers, confirming the role of reactive oxygen species in the observed immunomodulatory effects, independent of cytotoxicity. Furthermore, we report for the first time that, intranasal instillation of a single dose (5 mg/kg) of pristine or surfactant-dispersed ZnO NPs can cause pulmonary inflammation, already after 24 hr in a murine model. This was confirmed by up-regulation of eotaxin mRNA in the lung tissue and release of pro-inflammatory cytokines in the sera of mice exposed to ZnO NPs.
  
  
  CONCLUSIONS
  Our study highlights that even at sub-cytotoxic doses ZnO NPs can stimulate a strong inflammatory and antioxidant response in A549 cells. ZnO NP mediated cytotoxicity may be the outcome of failure of cellular redox machinery to contain excessive ROS formation. Moreover exposure to a single but relatively high dose of ZnO NPs via intranasal instillation may provoke acute pulmonary inflammatory reactions in vivo.
authors: Molecular Immunology Group, College of Public Health, Medical and Veterinary Sciences, Centre for Biodiscovery and Molecular Development of Therapeutics, James Cook University, Building 21, Molecular Sciences, James Cook Drive, Douglas Campus, Townsville, QLD, 4811, Australia. shruti.saptarshi@jcu.edu.au.
authors: Shruti R
authors: SR
authors: Saptarshi
authors: School of Medical Sciences, RMIT University, Melbourne, VIC, Australia. bryce.feltis@rmit.edu.au.|Department of Materials Engineering, Monash University, Melbourne, VIC, Australia. bryce.feltis@rmit.edu.au.
authors: Bryce N
authors: BN
authors: Feltis
authors: School of Medical Sciences, RMIT University, Melbourne, VIC, Australia. paul.wright@rmit.edu.au.
authors: Paul Fa
authors: PF
authors: Wright
authors: Molecular Immunology Group, College of Public Health, Medical and Veterinary Sciences, Centre for Biodiscovery and Molecular Development of Therapeutics, James Cook University, Building 21, Molecular Sciences, James Cook Drive, Douglas Campus, Townsville, QLD, 4811, Australia. andreas.lopata@jcu.edu.au.
authors: Andreas L
authors: AL
authors: Lopata
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology; D000281:Administration, Intranasal; D000818:Animals; D002460:Cell Line / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000187:drug effects / Q000276:immunology; D005260:Female; D051547:Heme Oxygenase-1 / Q000235:genetics; D006801:Humans; D007155:Immunologic Factors / Q000008:administration & dosage / Q000494:pharmacology*; D007249:Inflammation / Q000235:genetics; D016209:Interleukin-8 / Q000235:genetics; D020935:MAP Kinase Signaling System / Q000187:drug effects; D053768:Metal Nanoparticles / Q000008:administration & dosage* / Q000633:toxicity*; D008807:Mice, Inbred BALB C; D016328:NF-kappa B / Q000378:metabolism; D018675:Toxicity Tests / Q000379:methods; D015034:Zinc Oxide / Q000008:administration & dosage / Q000494:pharmacology*
chemical_list: D007155:Immunologic Factors; D016209:Interleukin-8; D016328:NF-kappa B; C496541:HMOX1 protein, human; D051547:Heme Oxygenase-1; D015034:Zinc Oxide; D000111:Acetylcysteine
references: Nanotoxicology. 2013 Jun;7(4):402-16
references: 22394310
references: ACS Nano. 2013 Dec 23;7(12):10621-35
references: 24187959
references: J Leukoc Biol. 2002 Nov;72(5):847-55
references: 12429706
references: Cell Biol Toxicol. 2014 Apr;30(2):79-88
references: 24554449
references: Chem Res Toxicol. 2012 Oct 15;25(10):2057-66
references: 22978249
references: Toxicol Appl Pharmacol. 2013 Jan 1;266(1):19-26
references: 23142470
references: Mol Cell Biochem. 1998 Nov;188(1-2):63-9
references: 9823012
references: J Nanobiotechnology. 2013;11:26
references: 23870291
references: J Appl Toxicol. 2013 Oct;33(10):1079-88
references: 23315988
references: PLoS One. 2013;8(6):e65704
references: 23755271
references: Nanoscale Res Lett. 2013 Nov 21;8(1):496
references: 24261419
references: Small. 2009 Apr;5(7):846-53
references: 19288475
references: Chem Biol Interact. 2012 Jun 25;198(1-3):29-37
references: 22640810
references: Nanotechnol Sci Appl. 2012 Aug 15;5:61-71
references: 24198497
references: Toxicol Sci. 2013 Nov;136(1):120-30
references: 23997113
references: Nanomedicine. 2011 Dec;7(6):904-13
references: 21664489
references: Chem Res Toxicol. 2008 Sep;21(9):1726-32
references: 18710264
references: J Mol Neurosci. 2012 Oct;48(2):464-71
references: 22528453
references: Science. 2006 Feb 3;311(5761):622-7
references: 16456071
references: J Appl Toxicol. 2009 Jan;29(1):69-78
references: 18756589
references: J Appl Toxicol. 2014 Apr;34(4):413-23
references: 24243578
references: Annu Rev Pharmacol Toxicol. 2010;50:323-54
references: 20055707
references: Toxicol Appl Pharmacol. 2012 Oct 1;264(1):94-103
references: 22842014
references: Toxicol Sci. 2012 Feb;125(2):462-72
references: 22112499
references: J Nutr. 2000 May;130(5S Suppl):1399S-406S
references: 10801951
references: Part Fibre Toxicol. 2011;8:27
references: 21896169
references: Nanotoxicology. 2012 Feb;6(1):22-35
references: 21332300
references: Part Fibre Toxicol. 2014;11:5
references: 24456852
references: Metallomics. 2011 Nov;3(11):1199-211
references: 21769377
references: Am J Respir Crit Care Med. 2010 Dec 1;182(11):1398-409
references: 20639441
references: Environ Health Perspect. 2007 Mar;115(3):403-9
references: 17431490
references: Drug Chem Toxicol. 2014 Jul;37(3):336-47
references: 24344737
references: Nanotoxicology. 2014 Sep;8(6):593-604
references: 23738974
references: Int J Mol Sci. 2013;14(8):15546-60
references: 23892599
references: Nanoscale. 2014 Jun 7;6(11):5791-8
references: 24740013
references: Toxicol In Vitro. 2013 Jun;27(4):1187-95
references: 23458966
references: Toxicol In Vitro. 2010 Feb;24(1):45-55
references: 19755143
references: Toxicol Appl Pharmacol. 2009 May 1;236(3):276-81
references: 19233222
references: Nanotoxicology. 2012 Nov;6(7):757-65
references: 22087559
references: Environ Mol Mutagen. 2011 Aug;52(7):582-9
references: 21786336
references: Part Fibre Toxicol. 2014;11:15
references: 24684892
references: Toxicol In Vitro. 2013 Mar;27(2):731-8
references: 23232460
references: Int J Nanomedicine. 2012;7:845-57
references: 22393286
references: ACS Nano. 2008 Oct 28;2(10):2121-34
references: 19206459
references: Environ Health Perspect. 2010 Jul;118(7):982-7
references: 20194077
references: Toxicol Lett. 2010 Dec 15;199(3):389-97
references: 20934491