(87g) Studies of Protein Expression Upon Cellular Exposure to Nanomaterials

Gabrielle Rogers-Nieman¹, David Lowry ²,

 Michael L. Kashon², Linda M. Sargent², and Cerasela Zoica Dinu¹*

¹Department of Chemical Engineering, West Virginia University, Morgantown WV,

26506, USA

²National Institute for Occupational Safety and Health, Morgantown WV, 26505, USA

* Corresponding author

Carbon nanotubes (CNTs) have many potential applications in industry because of their extraordinary mechanical, electrical and structural properties. However, recent studies demonstrate that CNTs can enter the working environment as suspended particulate matter of respirable sizes, which could pose an occupational inhalation exposure hazard and thus create challenges for exposure control. Research has also shown that CNTs associate with cellular elements, namely cytoskeletal filament microtubule and the DNA and interfere with cell progression to normal division, most notably by disruption of the centrosome. This leads to aneuploidy, an early event in the progression of lung mesothelioma. Such results further suggest that CNT cellular uptake and the mechanisms of nanotube cellular transport may play a role in both tumorigenesis and tumor progression. In our research we are unraveling the mechanisms associated with the nanotube uptake and their cellular transport in order to understand if the nanotube cellular interactions affect key protein expression involved in mitotic spindle formation. Using real time PCR complemented by optical microscopy and fluorescence activated cell sorting, our preliminary results show that CNTs affect the cell cycle and this is correlated with fluctuations in the expression level of integral motor proteins responsible for nanotube transport inside the cell. These observations are time and dose dependent. The quantification of expressional differences in messenger RNA of integral molecular motors during a time course of CNT exposure can provide a more detailed explanation of genotoxicity associated with cellular exposure to CNTs.

References:

Sargent LM, Hubbs AF, Young SH, Kashon ML, Dinu CZ, Salisbury JL, Benkovic SA, Lowry DT, Murray AR, Kisin ER, Siegrist KJ, Battelli L, Mastovich J, Sturgeon JL, Bunker KL, Shvedova AA, Reynolds SH. Single-walled carbon nanotube-induced mitotic disruption. Mutat Res. 2011. Dec 8. (Epub ahead of print)

Hubbs AF, Mercer RR, Benkovic SA, Harkema J, Sriram K, Schwegler-Berry D, Coravanahally MP, Nurkiwicz TR, Castranova V, Sargent LM. Nanotoxicology-a pathologist’s perspective. Toxicol Pathol. 2011 Feb; 39(2): 301-24/

Quintyne NJ, Schroer TA. Distinct cell cycle dependant roles for dynactin and dynein at centrosomes. J Cell Biol. 2002 Oct 28: 59(2): 245-54.

Kapitein LC, Peterman EJG, Kwok BH, Kim JH, Kapoor TM, Schmidt CF. The bipolar mitotic kinesin Eg5 moves on both microtubules that it crosslinks. Nature. 5 May 2005: 453, 114-118.

Gable A, Qiu M, Titus J, Balchand S, Fernez NP, Ma N, Collins ES, Fagerstrom C, Ross JL, Yang G, Wadsworth P. Dynamic reorganization of Eg5 in mammalian spindle throughout mitosis requires dynein and TPX2. Mol Biol Cell. 2012 Apr;23(7) 1254-66.

WiltshireC, SinghBL, Stockley J, Fleming J, Doyle B, Barnetson R, Robson CN, Kozielski F, Leung HY. Docetaxel-resistent Prostate cancer cells remain sensitive to S-trityl-L-cysteine-mediated Eg5 inhibition.  Mol Cancer Ther. 2010 Jun; 9(6): 1730-3.