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

作者：Stoehr Linda C; Endes Carola; Radauer Preiml Isabella; Boyles Matthew S P; Casals Eudald; Balog Sandor; Pesch Markus; Petri Fink Alke; Rothen Rutishauser Barbara; Himly Martin; Clift Martin J D; Duschl Albert^*

来源：Particle and Fibre Toxicology, 2015, 12(1): 29.

DOI：10.1186/s12989-015-0104-6

• A549 cells
• Interleukin-8
• Air-liquid interface
• Submerged cultures
• Acute pulmonary (pro-)inflammatory effects

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 mu g/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-alpha treatment. Both pIL8-GFP and pIL8-RFP also showed a marked signal induction in response to TNF-alpha, 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.

• 出版日期2015-9-29