Evaluation of the cytotoxic, genotoxic and inflammatory effects induced by the exposure to mineral fibres in in vitro human cellular models

Exposure to mineral fibres represents a serious occupational and environmental hazard, since it leads to chronic lung inflammation with the subsequent development of pneumoconiosis, fibrotic pulmonary diseases, lung cancer and malignant mesothelioma. However, since the toxicity and carcinogenicity of mineral fibres is profoundly interwoven with a broad variety of crystal-chemical-physical characteristics, the biogeochemical mechanisms through which mineral fibres, and especially asbestos, induce adverse effects in vivo are yet to be completely understood. The first part of this study aimed at investigating the different possible toxicity mechanisms of UICC standard crocidolite, chrysotile from Balangero and erionite from Jersey, which are respectively representative of the classes of amphibole asbestos, serpentine asbestos and fibrous erionite. The acute damaging effects exerted by these mineral fibres during the first 24 h of exposure were evaluated by performing a comparative study in an in vitro THP-1 cellular model of M0, M1 and M2 macrophages. The toxicity mechanisms of the three mineral fibres appeared to differ significantly. Crocidolite seemed to exert its toxic effects mostly as a result of its biodurability, ROS production, cytokine release and DNA damage. Chrysotile, due to its low biodurability, displayed toxic effects correlated with the release of toxic metals and the production of ROS and cytokines. Other mechanisms were involved in explaining the toxicity of biodurable fibrous erionite, which induced lower ROS and toxic metal release, but exhibited a cation exchange capacity able to alter the intracellular homeostasis of important cations. Interestingly, M2 macrophages, which are characteristically known for countering the inflammatory response, in the presence of asbestos fibres and erionite, exacerbated this process by secreting pro-inflammatory mediators. For the second part of this project, representative batches of short (length < 5 µm) and long (length > 5 µm) chrysotile fibres were prepared by cryogenic milling of a commercial chrysotile extracted from an open pit mine near Yasny (Russia). To gain new insights into the toxicity mechanisms of these size-separated fractions of chrysotile, their cytotoxic and genotoxic activity was investigated in an in vitro cellular model based on human THP-1-derived M0 macrophages and HECV endothelial cells, both separately as well as in a co-culture setup. At all time-points, both chrysotile fractions displayed significant acute cytotoxic effects, with results that were comparable to the well-known damaging effects of crocidolite. In particular, the long fraction (> 5 μm) of chrysotile showed a notably higher cytotoxic potential, causing ROS production, DNA damage and the transcriptional upregulation of inflammatory mediators in M0 macrophages. On the other hand, the short fraction of chrysotile displayed a significant degree of cytotoxicity and genotoxicity in vitro, which is quite intriguing considering the ongoing debate regarding the potential toxicity of short asbestos fibres (< 5 μm). It should also be noted that in HECV cells the mineral fibres caused toxicity mainly as a result of direct cell membrane damage, whereas in M0 macrophages the apoptotic mechanism had a fundamental role in the induction of cell death.