Inflammatory and carcinogenic potential of size-separated chrysotile fibres assessed through in vitro models of human lung tissue

Asbestos minerals have been widely exploited due to their physical-chemical properties, and chrysotile asbestos has accounted for about 95% of all asbestos commercially employed worldwide. The exposure to chrysotile, classified like other five amphibole asbestos species as carcinogenic to humans, represents a serious occupational and environmental hazard. Nevertheless, this mineral is still largely employed in about 65% of the countries worldwide, which still allow its “safe use”.

The complex mechanisms through which the mineral fibres induce toxicity are not yet completely understood. In this regard, the morphometric parameters of asbestos fibres (e.g., length, width, aspect ratio) are known for their fundamental role in determining the degree of pathogenicity. Thus, the potential toxicity of short chrysotile fibres remains widely debated due to the contradictory results from countless studies. The present study investigated the different toxicity mechanisms of two representative batches of short (length <5 µm) and long (length >5 µm) chrysotile fibres obtained by cryogenic milling. The cytotoxic, genotoxic, and pro-inflammatory potential of the two chrysotile fractions, as compared to crocidolite and wollastonite carcinogenic positive and negative controls, was investigated on human THP-1-derived macrophages and HECV endothelial cells, both separately and in a co-culture setup, mimicking the alveolar pro-inflammatory microenvironment, in time course experiments up to 1 week. Parallel exposure experiments, up to 12 days, were also run on an in vitro 3D tissue model, the Mattek EpiAirway™, closely resembling the physiology of the mature human bronchial epithelium. Through these models, we could assess that both chrysotile fractions displayed cytotoxic, genotoxic, and pro-inflammatory effects, with resulted comparable to the well-known damaging effects of crocidolite asbestos, or higher, as in the case of the longer chrysotile fraction. Furthermore, in presence of HECV, fibre-treated macrophages showed prolonged inflammation, indicating an interesting crosstalk between these cells, able to sustain a low-grade chronic inflammation in the lung. This was also confirmed in the 3D lung tissue model were a semi-chronic exposure of 12 days led to a prolonged inflammatory response in crocidolite- and chrysotile-treated tissues as compared to control, untreated ones. In conclusion, these results help to shed light on some important open questions on the mechanisms of toxicity of chrysotile asbestos fibres.

 

Acknowledgements

This project has received funding from the Italian Ministry of University and Research, PRIN-2017 “FIBRES”, and from the Italian Ministry of Health for “Research and development of projects of alternative methods to animal models through experimental technologies, 2022”.