Abstract
Agglomeration and sedimentation of nanoparticles (NPs) within biological solutions is a major limitation in their use in many downstream applications. It has been proposed that serum proteins associate with the NP surface to form a protein corona that limits agglomeration and sedimentation. Here, we investigate the effect of fetal bovine serum (FBS) proteins on the dispersion stability, dosimetry, and NP-induced cytotoxicity of cationic zinc oxide nanoparticles (nZnO) synthesized via forced hydrolysis with a core size of 10 nm. Two different in vitro cell culture models, suspension and adherent, were evaluated by comparing a phosphate buffered saline (PBS) nZnO dispersion (nZnO/PBS) and an FBS-stabilized PBS nZnO dispersion (nZnO – FBS/PBS). Surface interactions of FBS on nZnO were analyzed via spectroscopic and optical techniques. Fourier transformed infrared spectroscopy (FTIR) confirmed the adsorption of negatively charged protein components on the cationic nZnO surface through the disappearance of surfaced-adsorbed carboxyl functional groups and the subsequent detection of vibrational modes associated with the protein backbone of FBS-associated proteins. Further confirmation of these interactions was noted in the isoelectric point shift of the nZnO from the characteristic pH of 9.5 to a pH of 6.1. In nZnO – FBS/PBS dispersions, the FBS reduced agglomeration and sedimentation behaviors to impart long-term improvements (>24 h) to the nZnO dispersion stability. Furthermore, mathematical dosimetry models indicate that nZnO – FBS/PBS dispersions had consistent NP deposition patterns over time unlike unstable nZnO/PBS dispersions. In suspension cell models, the stable nZnO – FBS/PBS dispersion resulted in a ~33 % increase in the NP-induced cytotoxicity for both Jurkat leukemic and Hut-78 lymphoma cancer cells. In contrast, the nZnO – FBS/PBS dispersion resulted in 49 and 71 % reductions in the cytotoxicity observed towards the adherent breast (T-47D) and prostate (LNCaP) cancer cell lines, respectively. Presence of FBS in the NP dispersions also increased the reactive oxygen species generation. These observations indicate that the improved dispersion stability leads to increased NP bioavailability for suspension cell models and reduced NP sedimentation onto adherent cell layers resulting in more accurate in vitro toxicity assessments.
Highlights
When prepared in nanostructured form, many materials develop novel and unique physicochemical properties leading to their use in nanotechnology applications such as quantum dots and MRI contrast agents for imaging [1], biosensors that use surface plasmon resonance [2], and nanomedicinal approaches that employ hyperthermia and photothermal therapy [3]
The positive to negative shift in the zeta potential observed for the NP-induced cytotoxicity of cationic zinc oxide nanoparticles (nZnO) – FBSdried NPs is indicative of the absorption of negatively charged protein components on the surface of the nZnO [51]
The IC50 value for the Hut cells (Fig. 11c) showed a decrease from 0.21 ± 0.03 mM when treated with the nZnO/phosphate buffered saline (PBS) dispersion to 0.14 ± 0.02 mM when treated with the nZnO – fetal bovine serum (FBS)/PBS dispersion. These results clearly demonstrate that coating nZnO with FBS proteins and the resulting improved dispersion stability increases their toxic response to cancer cells by ~33 % for both cell types
Summary
When prepared in nanostructured form, many materials develop novel and unique physicochemical properties leading to their use in nanotechnology applications such as quantum dots and MRI contrast agents for imaging [1], biosensors that use surface plasmon resonance [2], and nanomedicinal approaches that employ hyperthermia and photothermal therapy [3]. Cytotoxicity is a property that is inherent in many of these commonly used materials when they are synthesized with their crystallite size in the nanoscale range [7,8,9] This has raised significant scientific and public concerns on the environmental and health effects of every nanomaterial before employing them in nanotechnology applications. Some studies have shown that the toxic response of a given nanomaterial might vary with the specific cell/organism type such as prokaryotes versus eukaryotes [15], gram-positive bacteria versus gram-negative bacteria, and normal cells versus cancerous cells [16]. Toxicity studies are extremely important both to assess the environmental and health impacts of NPs and to modify the nanomaterial design to make them safer, as well as to develop novel nanomedicinal approaches utilizing cell-specific cytotoxicity
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