M.E. Johnson, A.R. Montoro Bustos, S.K. Hanna, R. Dave Holbrook, C.M.S. Ims, P.E. Lapasset, E.J. Petersen, J.T. Elliot, B.C. Nelson, L.L. Yu, K.E. Murphy, M.R. Winchester
National Institute of Standards and Technology,
Keywords: nanoparticles, AuNPs, Caenorhabditis elegans, spICP-MS, SEM/EDX
Summary:The prevalent use and incorporation of engineered nanoparticles in consumer products increases the likelihood of their interaction with environmental and biological systems, and thus, has fostered research efforts to understand their potential risks and implications. Caenorhabditis elegans (C. elegans) is an ideal model organism for nanotoxicity studies because of its short life span, ease of culturing, and transparency, allowing for visualization of changes in nematode morphology in the presence of toxicants. Several research efforts have focused on uptake and effects of gold nanoparticles (AuNPs) on C. elegans. Ingestion of AuNPs, inhibition of reproduction, and maternal transfer were among some of the primary findings. Here we present results of size dependent uptake of AuNPs by C. elegans. Nematodes were exposed to 80 nm, 100 nm, and 150 nm citrate-stabilized AuNPs for 24 h at equal Au mass fraction concentrations (Figure 1 and Table 1). Common washing procedures were previously determined to be insufficient in removing excess suspended AuNPs after exposure, therefore, a sucrose density gradient centrifugation protocol (Figure 1A) was employed to separate the nematodes from AuNPs freely suspended in the exposure media.3 Image analysis of samples in all exposure conditions by scanning electron microscopy combined with energy dispersive X-ray spectroscopy (SEM/EDX) was performed to assess the efficacy of the C. elegans/AuNP separation. Quantification of total Au uptake in dry samples of non-exposed and Au exposed C. elegans was determined following acid digestions by conventional inductively coupled plasma mass spectrometry (ICP-MS) analyses. Alternatively, alkaline digestions of biological tissue were utilized for particle sizing in single particle ICP-MS analyses. Nanoparticulate body burdens were measured for each AuNP uptake condition. Finally, size distributions and particle number concentrations were determined for all AuNPs dispersed in water and AuNPs taken up by C. elegans. ICP-MS results suggest a size dependent AuNP uptake. Under these conditions, we found an increase in total Au uptake (related to Au concentration) for the 80 nm and 100 nm AuNPs, however there was reduced uptake for the 150 nm AuNP exposure. This reduction may be due to the decreases in total number of particles in solution with increased particle size. The analysis of the liberated AuNPs in C. elegans revealed an alteration in the size distribution of NPs, which may be a result of the NPs being exposed to physiological processes (i.e. gut compaction and/or digestion) within the nematode.