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Current Topics in Biochemical Research   Volumes    Volume 14  Issue 1
Mass spectrometry-based proteomic assessment of the in vitro toxicity of carbon nanotubes
P. Kumarathasan, D. Das, M. A. Salam, S. Mohottalage, N. DeSilva, B. Simard, R. Vincent
Pages: 15 - 27
Number of pages: 13
Current Topics in Biochemical Research
Volume 14  Issue 1

Copyright © 2012 Research Trends. All rights reserved

Due to the unique size-specific properties, carbon nanotubes (CNTs) have been incorporated in several industrial processes and consumer products, thereby increasing the likelihood of some human exposure. However, the few studies on toxicity of CNTs in the literature are often not comparable, due in part to lack of information on physicochemical characteristics. We have compared the biological responses of J774 murine macrophages exposed to four well characterized nanomaterials: single-walled and multi-walled, pristine and oxidatively-modified CNTs, at a dose range of 0-100 μg/cm2. Viability was assessed by AlamarBlue and MTS reduction. Cell supernatants were analyzed for oxidatively modified protein metabolites by HPLC-coulometric array detection. Shot-gun proteomic analyses of direct and tryptic-digested cell lysates were performed by MALDI-TOF-TOF-MS. Mass spectral profiles were interrogated in the m/z region <6 kDa using k-nearest neighbour clustering algorithm. All CNTs were cytotoxic. J774 cells exhibited mass spectral patterns specific to the CNT exposures. Data-mining revealed an elevation in cellular endothelin-1, a pro-inflammatory and mitogenic peptide, and a decrease in cytoplasmic lactate dehydrogenase levels, an indicator of cell membrane permeability, notably in response to the oxidized CNTs. Elevation of o-tyrosine, consistent with formation of reactive oxygen species, was coherent with proteomic changes. The increased potency can be attributed to surface modifications and structural changes in oxidized CNTs, namely increased specific surface area and pore size, and presence of carboxylic groups. Our observations under specific in vitro conditions indicate that surface functionalities, and not the metal contaminants, are driving the biological reactivity of these CNTs.
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