Comparative characteristics of various fibrous materials in in vitro experiments

Aim. Comparative assessment of the effect of fibrous materials on cell cultures RAW264.7 and BEAS-2B. Methods. The effects of various fibrous materials single-walled carbon nanotubes of two types (SWCNT-1 and SWCNT-2), differing in morphological characteristics, and chrysotile asbestos as a positive control was assessed on two cell lines macrophages RAW 264.7 and human bronchial epithelium BEAS-2B cells. The studied materials concentration range for experiments on cells was selected taking into account the SWCNT content in the air of the working area and the subsequent modeling of SWCNT deposition in the human respiratory tract. Suspensions of the studied materials were prepared based on cell culture media by ultrasonication. Cytotoxicity assessment after 48 hours of incubation was performed by using the MTS colorimetric assay. The expression level of apoptosis markers was assessed by immunoblotting using the corresponding monoclonal antibodies. Visualization of SWCNT-1, SWCNT-2 and chrysotile asbestos in BEAS-2B cell cultures was carried out by improved dark-field microscopy. Results. According to dark-field microscopy, all the studied fibrous materials were found on the surface or cytoplasm of the cells. SWCNT and chrysotile asbestos did not have a direct cytotoxic effect in the MTS assay and did not induce apoptosis according to the results of Western blotting in cell cultures of RAW264.7 macrophages and BEAS-2B bronchial epithelium. In the cells of the bronchial epithelium (BEAS-2B) that showed greater sensitivity, a slight increase in the expression of pro-apoptotic protein PARP, which was more pronounced for shorter SWCNT-2, was revealed. Conclusion. Both types of SWCNTs, despite the differences in morphological characteristics, demonstrated similar effects in in vitro experiments; this result, with its further verification, can have an important practical application in justifying approaches to determining the safety criteria for single-walled carbon nanotubes as a class of nanomaterials of the same type.

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Faculty Opinions recommendation of In vitro toxicity evaluation of single walled carbon nanotubes on human A549 lung cells.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1083823.536755 ◽

2007 ◽

Author(s):

Ruth Roberts

Keyword(s):

Carbon Nanotubes ◽

Single Walled Carbon Nanotubes ◽

In Vitro Toxicity ◽

Lung Cells ◽

Toxicity Evaluation ◽

Single Walled Carbon ◽

Walled Carbon Nanotubes ◽

Human A549 ◽

A549 Lung

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New Mechanistic Insights on Carbon Nanotubes’ Nanotoxicity Using Isolated Submitochondrial Particles, Molecular Docking, and Nano-QSTR Approaches

Biology ◽

10.3390/biology10030171 ◽

2021 ◽

Vol 10 (3) ◽

pp. 171

Author(s):

Michael González-Durruthy ◽

Riccardo Concu ◽

Juan M. Ruso ◽

M. Natália D. S. Cordeiro

Keyword(s):

Carbon Nanotubes ◽

Network Models ◽

Single Walled Carbon Nanotubes ◽

Docking Studies ◽

Fractal Surface ◽

Submitochondrial Particles ◽

Elastic Network ◽

Elastic Network Models ◽

Walled Carbon Nanotubes

Single-walled carbon nanotubes can induce mitochondrial F0F1-ATPase nanotoxicity through inhibition. To completely characterize the mechanistic effect triggering the toxicity, we have developed a new approach based on the combination of experimental and computational study, since the use of only one or few techniques may not fully describe the phenomena. To this end, the in vitro inhibition responses in submitochondrial particles (SMP) was combined with docking, elastic network models, fractal surface analysis, and Nano-QSTR models. In vitro studies suggest that inhibition responses in SMP of F0F1-ATPase enzyme were strongly dependent on the concentration assay (from 3 to 5 µg/mL) for both pristine and COOH single-walled carbon nanotubes types (SWCNT). Besides, both SWCNTs show an interaction inhibition pattern mimicking the oligomycin A (the specific mitochondria F0F1-ATPase inhibitor blocking the c-ring F0 subunit). Performed docking studies denote the best crystallography binding pose obtained for the docking complexes based on the free energy of binding (FEB) fit well with the in vitro evidence from the thermodynamics point of view, following an affinity order such as: FEB (oligomycin A/F0-ATPase complex) = −9.8 kcal/mol > FEB (SWCNT-COOH/F0-ATPase complex) = −6.8 kcal/mol ~ FEB (SWCNT-pristine complex) = −5.9 kcal/mol, with predominance of van der Waals hydrophobic nano-interactions with key F0-ATPase binding site residues (Phe 55 and Phe 64). Elastic network models and fractal surface analysis were performed to study conformational perturbations induced by SWCNT. Our results suggest that interaction may be triggering abnormal allosteric responses and signals propagation in the inter-residue network, which could affect the substrate recognition ligand geometrical specificity of the F0F1-ATPase enzyme in order (SWCNT-pristine > SWCNT-COOH). In addition, Nano-QSTR models have been developed to predict toxicity induced by both SWCNTs, using results of in vitro and docking studies. Results show that this method may be used for the fast prediction of the nanotoxicity induced by SWCNT, avoiding time- and money-consuming techniques. Overall, the obtained results may open new avenues toward to the better understanding and prediction of new nanotoxicity mechanisms, rational drug design-based nanotechnology, and potential biomedical application in precision nanomedicine.

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