Adverse effects of long and straight multiwalled carbon nanotubes on cytoskeleton, nuclear lamina, and genome-wide transcriptome of primary human mesothelial cells

Question Certain long and straight multiwalled carbon nanotubes (MWCNTs) have shown carcinogenic potential in animal models after intraperitoneal application. Nonetheless, the underlying mechanisms are still unclear. Owing to their fiber-like structure, the long and straight MWCNTs seem to cause more adverse effects than e.g. tangled types. These effects include the potential to cause malignant mesothelioma, a cancer originating from mesothelial cells, and often associated with asbestos exposure. Methods To gain insights into pertinent disease mechanisms, we analyzed the biological effects of two tailor-made MWCNTs (CNT2: slightly curved, diameter 0.050 μm, length 4.18 μm; CNT 6: straight, diameter 0.049 μm, length 10.9 μm), and a reference MWCNT (JRC NM-401: straight, diameter 0.067 μm, length 4.04 μm). These MWCNTs were tested for sterility and endotoxin content, and were free of relevant contamination. As a disease-relevant cell model, we used primary human peritoneal mesothelial cells (LP9) treated as submerged cultures for 24 h with 5 μg/cm2 of the different fibers. Results Immunofluorescence of alpha-tubulin, a protein that forms part of microtubules (a major cytoskeleton component), showed extensive abnormalities. Similarly, we observed depletion of lamin B1 (LMNB1), which is one of the structural proteins of the nuclear lamina and important in the maintenance of nuclear structure and function. After microarray analysis, we found many differentially-expressed genes relative to untreated control, i.e. 7609, 7478, 5742 for NM-401, CNT2, and CNT6, respectively. Bioinformatic analyses revealed many common significantly activated pathways and molecular functions, associated with inflammation and carcinogenesis, notably lung cancer. The top activated canonical pathways include IL6, IL8, and p38 MAPK signaling. Conclusion In conclusion, our results show that exposure to certain long and straight MWCNTs can lead to marked disturbance of cellular structure and the transcriptome of mesothelial cells, eventually contributing to carcinogenesis