title: Assessment of a panel of interleukin-8 reporter lung epithelial cell lines to monitor the pro-inflammatory response following zinc oxide nanoparticle exposure under different cell culture conditions.
abstract: BACKGROUND
  Stably transfected lung epithelial reporter cell lines pose an advantageous alternative to replace complex experimental techniques to monitor the pro-inflammatory response following nanoparticle (NP) exposure. Previously, reporter cell lines have been used under submerged culture conditions, however, their potential usefulness in combination with air-liquid interface (ALI) exposures is currently unknown. Therefore, the aim of the present study was to compare a panel of interleukin-8 promoter (pIL8)-reporter cell lines (i.e. green or red fluorescent protein (GFP, RFP), and luciferase (Luc)), originating from A549 lung epithelial type II-like cells cells, following NPs exposure under both submerged and ALI conditions.
  
  
  METHODS
  All cell lines were exposed to zinc oxide (ZnO) NPs at 0.6 and 6.2 μg/cm(2) for 3 and 16 hours under both submerged and ALI conditions. Following physicochemical characterization, the cytotoxic profile of the ZnO-NPs was determined for each exposure scenario. Expression of IL-8 from all cell types was analyzed at the promoter level and compared to the mRNA (qRT-PCR) and protein level (ELISA).
  
  
  RESULTS
  In summary, each reporter cell line detected acute pro-inflammatory effects following ZnO exposure under each condition tested. The pIL8-Luc cell line was the most sensitive in terms of reporter signal strength and onset velocity following TNF-α treatment. Both pIL8-GFP and pIL8-RFP also showed a marked signal induction in response to TNF-α, although only after 16 hrs. In terms of ZnO-NP-induced cytotoxicity pIL8-RFP cells were the most affected, whilst the pIL8-Luc were found the least responsive.
  
  
  CONCLUSIONS
  In conclusion, the use of fluorescence-based reporter cell lines can provide a useful tool in screening the pro-inflammatory response following NP exposure in both submerged and ALI cell cultures.
authors.affiliation: Department of Molecular Biology, University of Salzburg, Hellbrunnerstrasse 34, 5020, Salzburg, Austria. lindacorinna.stoehr@stud.sbg.ac.at.|Grimm Aerosol Technik GmbH & Co. KG, Ainring, Germany. lindacorinna.stoehr@stud.sbg.ac.at.
authors.forename: Linda C
authors.initials: LC
authors.lastname: Stoehr
authors.affiliation: BioNanomaterials, Adolphe Merkle Institute, Université de Fribourg, Fribourg, Switzerland. carola.endes@unifr.ch.
authors.forename: Carola
authors.initials: C
authors.lastname: Endes
authors.affiliation: Department of Molecular Biology, University of Salzburg, Hellbrunnerstrasse 34, 5020, Salzburg, Austria. isabella.radauer-preiml@sbg.ac.at.
authors.forename: Isabella
authors.initials: I
authors.lastname: Radauer-Preiml
authors.affiliation: Department of Molecular Biology, University of Salzburg, Hellbrunnerstrasse 34, 5020, Salzburg, Austria. matthew.boyles@sbg.ac.at.
authors.forename: Matthew S P
authors.initials: MS
authors.lastname: Boyles
authors.affiliation: Institut Català de Nanotecnologia (ICN), Bellaterra, Spain. eudald.casals@icn.cat.
authors.forename: Eudald
authors.initials: E
authors.lastname: Casals
authors.affiliation: Soft Matter Scattering, Adolphe Merkle Institute, Université de Fribourg, Fribourg, Switzerland. sandor.balog@unifr.ch.
authors.forename: Sandor
authors.initials: S
authors.lastname: Balog
authors.affiliation: Grimm Aerosol Technik GmbH & Co. KG, Ainring, Germany. mp@grimm-aerosol.com.
authors.forename: Markus
authors.initials: M
authors.lastname: Pesch
authors.affiliation: BioNanomaterials, Adolphe Merkle Institute, Université de Fribourg, Fribourg, Switzerland. alke.fink@unifr.ch.
authors.forename: Alke
authors.initials: A
authors.lastname: Petri-Fink
authors.affiliation: BioNanomaterials, Adolphe Merkle Institute, Université de Fribourg, Fribourg, Switzerland. barbara.rothen@unifr.ch.
authors.forename: Barbara
authors.initials: B
authors.lastname: Rothen-Rutishauser
authors.affiliation: Department of Molecular Biology, University of Salzburg, Hellbrunnerstrasse 34, 5020, Salzburg, Austria. martin.himly@sbg.sc.at.
authors.forename: Martin
authors.initials: M
authors.lastname: Himly
authors.affiliation: BioNanomaterials, Adolphe Merkle Institute, Université de Fribourg, Fribourg, Switzerland. martin.clift@unifr.ch.
authors.forename: Martin J D
authors.initials: MJ
authors.lastname: Clift
authors.affiliation: Department of Molecular Biology, University of Salzburg, Hellbrunnerstrasse 34, 5020, Salzburg, Austria. albert.duschl@sbg.ac.at.
authors.forename: Albert
authors.initials: A
authors.lastname: Duschl
mesh_terms: D002460:Cell Line; D004847:Epithelial Cells / Q000378:metabolism; D017930:Genes, Reporter; D006801:Humans; D007249:Inflammation / Q000139:chemically induced* / Q000378:metabolism; D016209:Interleukin-8 / Q000235:genetics*; D008168:Lung / Q000166:cytology / Q000378:metabolism*; D053768:Metal Nanoparticles / Q000633:toxicity*; D015034:Zinc Oxide / Q000633:toxicity*
chemical_list: D016209:Interleukin-8; D015034:Zinc Oxide