An alternative in vitro model considering cell-cell interactions in fiber-induced pulmonary fibrosis

Abstract

Particularly since the wide-ranging health effects of asbestos exposure became known, great emphasis has been placed on detailed toxicity testing of known but also newly developed fiber materials. Exposure to respirable pollutants like fibers can lead to tissue injury causing lung diseases such as pulmonary fibrosis or cancer. In order to detect the toxic potential of such aerosols at an early stage, the development of suitable test systems is essential. In this study, we illustrate the development of an advanced in vitro cell model closely resembling the physiological structure of the alveoli, and we highlight its advantages over simpler models to predict pro-fibrotic changes. For this reason, we analyzed the cytotoxic effects of fiber-like multi-walled carbon nanotubes after 24 and 48 h exposure, and we investigated inflammatory, genotoxic and pro-fibrotic changes occurring in the developed triple culture consisting of lung epithelial cells, macrophages and fibroblasts compared to a co-culture of epithelial cells and fibroblasts or a mono culture of epithelial cells. In summary, the triple culture system is more precisely able to detect a pro-fibrotic phenotype including epithelial-mesenchymal transition as well as secondary genotoxicity, even if exhibiting lower cytotoxicity in contrast to the less advanced systems. These effects might be traced back to the complex interplay between the different cell types, all of which play an important role in the inflammatory response, which precedes wound healing, or even fibrosis or cancer development.

Keywords:

• MWCNTs
• oxidative stress
• inflammation
• secondary genotoxicity
• EMT

Acknowledgements

The authors thank Dr. Armin Springer (Elektronenmikroskopisches Zentrum (EMZ), University of Rostock) as executive and Arne Koch for the organization and support (Joint Mass Spectrometry Center at Analytical Chemistry, Institute of Chemistry, University of Rostock), as well as Dr. Marcus Frank (EMZ) for providing the relevant equipment for the SEM and EDX analysis. Furthermore, The authors thank Dr. Yaobo Ding (Institute of Lung Health and Immunity (LHI) and the Comprehensive Pneumology Center (CPC), Helmholtz Zentrum München) for the support of the DLS measurements as well as Dr. Tobias Stöger (LHI/CPC) for providing the relevant equipment.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Data availability statement

The data that support the findings of this study are available from the corresponding author, upon request.

Additional information

Funding

This work was co-financed by the European Union (European Fund for regional development) via the Technologie-Beratungs-Institut GmbH (TBI, Technology Consulting Institute of the Ministry of Economic Affairs, Infrastructure, Tourism and Labor Mecklenburg-Western Pomerania, project number TBI-V-1-262-VBW-093). It was supported by the Bayerische Staatsministerium für Umwelt und Verbraucherschutz (Bavarian State Ministry for the Environment and Consumer Protection, project number TLK01L-77228) in the frame of “Biologische Antwort auf Partikel in einem Lungenmodel” (Biological response to particles in a lung model) as well as by the Biogenic Organotropic Wetsuits (BOW) project funded by the European Union’s Horizon 2020 research and innovation program under grant agreement number 952183.