Download A newly developed in vitro model of the human epithelial airway

Air-liquid exposure of gold nanoparticles to study effects, uptake and
intracellular distribution in a human 3D epithelial airway model
C. Brandenberger1, A. G. Lenz2, C. Mühlfeld1,3, O. Schmid2, P. Gehr1 and B. RothenRutishauser1
1
Institute of Anatomy, Division of Histology, University of Bern, Bern, Switzerland
Institute of Lung Biology and Disease, Helmholtz Zentrum München, Germany
3
Institute of Anatomy and Cell Biology, Justus-Liebig-University Giessen, Germany
2
Nanoparticles (NPs <100nm) show promising features for pharmaceutical applications. However,
biomedical applications of NPs require a detailed understanding of potential interactions with
biological systems. One important route of entry into the body is pulmonary inhalation, which
could potentially be used for biomedical applications. To analyse particle-cell interactions
within in vitro cell studies, a dose-controlled system for delivery of NPs to lung cell cultures
is required. A new air-liquid interface cell exposure (ALICE) system (Fig. 1) has therefore
been developed, allowing a dose-controlled deposition of NPs on top of the cells [1]. This
system has been shown to mimic the aerosol inhalation very realistically.
The ALICE system was combined with either a human triple cell co-culture system,
composed of epithelial cells, macrophages and dendritic cells, simulating the most important
barrier functions of the lung epithelial airway [2], or with epithelial mono-cultures only. The
cells were exposed to 15nm gold particles, which show promising features for biomedical
applications. Oxidative stress and pro-inflammatory effects were analyzed at concentrations
of 61ng Au/cm2 and 561ng Au/cm2 deposition but no adverse effects were found after 4h and
24h exposure time [3].
The intracellular particle distribution overtime was quantitatively evaluated by stereology on
electron microscopic images and compared to particle uptake under submerged conditions.
The analysis revealed a significant, non-random intracellular NP distribution. NPs were
localized in intracellular vesicles, but not in the nucleus, mitochondria, endoplasmatic
reticulum or golgi. Only a minority of gold NPs was found free in the cytosol. However,
when NPs were exposed under submerged conditions, an increased NP agglomeration was
observed. This can lead to differences in cellular NP uptake mechanisms, hence influence
particle-cell interactions. In addition, it was found that particle-cell interaction was reduced
under submerged exposure conditions, since not all NPs suspended in the cell culture medium
were able to deposit on the cells.
With the ALICE system, it is therefore possible to reflect the physiological conditions of
aerosol inhalation and deposition more precisely. No adverse effects from gold NPs were
observed. By using a stereological analysis the intracellular particle number, localisation and
intracellular trafficking can be quantified.
[1] Lenz et al. Part Fibre Toxicol 6:32 (2009).
[2] Rothen-Rutishauser et al. Exp Opin Drug Metabol Toxicol 4(8):1075-1089 (2008).
[3] Brandenberger et al. Toxicol Appl Pharmacol 242(1):56-65 (2010).
This work was supported by the Doerenkamp Zbinden Foundation and AnimalFreeResearch.