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Nanodiamonds act as Trojan horse for intracellular delivery of metal ions to trigger cytotoxicity.

Zhu Y, Zhang Y, Shi G, Yang J, Zhang J, Li W, Li A, Tai R, Fang H, Fan C, Huang Q - Part Fibre Toxicol (2015)

Bottom Line: In addition, theoretical calculation and molecular dynamics (MD) computation were used to illustrate the adsorption properties of different metal ion on NDs as well as release profile of ion from ND-ion complexes at different pH values.Detailed investigation of ND-Cu2+ interaction showed that the amount of released Cu2+ from ND-Cu2+ complexes at acidic lysosomal conditions was much higher than that at neutral conditions, leading to the elevation of intracellular ROS level, which triggered cytotoxicity.The present experimental and theoretical results provide useful insight into understanding of cytotoxicity triggered by nanoparticle-ion interactions, and open new ways in the interpretation of nanotoxicity.

View Article: PubMed Central - PubMed

Affiliation: Division of Physical Biology, and Bioimaging Center, Shanghai Synchrotron Radiation Facility, Shanghai Institute of Applied Physics, Chinese Academy of Sciences, Shanghai, 201800, China. zhuying@sinap.ac.cn.

ABSTRACT

Background: Nanomaterials hold great promise for applications in the delivery of various molecules with poor cell penetration, yet its potential for delivery of metal ions is rarely considered. Particularly, there is limited insight about the cytotoxicity triggered by nanoparticle-ion interactions. Oxidative stress is one of the major toxicological mechanisms for nanomaterials, and we propose that it may also contribute to nanoparticle-ion complexes induced cytotoxicity.

Methods: To explore the potential of nanodiamonds (NDs) as vehicles for metal ion delivery, we used a broad range of experimental techniques that aimed at getting a comprehensive assessment of cell responses after exposure of NDs, metal ions, or ND-ion mixture: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, Trypan blue exclusion text, optical microscope observation, synchrotron-based scanning transmission X-ray microscopy (STXM) and micro X-ray fluorescence (μXRF) microscopy, inductively coupled plasma-mass spectrometry (ICP-MS), reactive oxygen species (ROS) assay and transmission electron microscopy (TEM) observation. In addition, theoretical calculation and molecular dynamics (MD) computation were used to illustrate the adsorption properties of different metal ion on NDs as well as release profile of ion from ND-ion complexes at different pH values.

Results: The adsorption capacity of NDs for different metal ions was different, and the adsorption for Cu2+ was the most strong among divalent metal ions. These different ND-ion complexes then had different cytotoxicity by influencing the subsequent cellular responses. Detailed investigation of ND-Cu2+ interaction showed that the amount of released Cu2+ from ND-Cu2+ complexes at acidic lysosomal conditions was much higher than that at neutral conditions, leading to the elevation of intracellular ROS level, which triggered cytotoxicity. By theoretical approaches, we demonstrated that the functional carbon surface and cluster structures of NDs made them good vehicles for metal ions delivery.

Conclusions: NDs played the Trojan horse role by allowing large amounts of metal ions accumulate into living cells followed by subsequent release of ions in the interior of cells, which then led to cytotoxicity. The present experimental and theoretical results provide useful insight into understanding of cytotoxicity triggered by nanoparticle-ion interactions, and open new ways in the interpretation of nanotoxicity.

No MeSH data available.


Related in: MedlinePlus

Enhancement of bioeffects by ND-vectorized Cu2+. a and b: Enhancement of Cellular ROS level by ND-vectorized Cu2+. a: ROS generation, b: NAC protection (*p < 0.05, **p < 0.01, one-way ANOVA for comparison). c: TEM images of a typical L929 cell after incubation with NDs, Cu2+ and NDs-Cu2+ mixture for 24 h. Arrows indicate NDs (Scale bars = 2 μm). d: Flow cytometric analysis of L929 cells after incubation with NDs, Cu2+ and NDs-Cu2+ mixture for 24 h.
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Fig5: Enhancement of bioeffects by ND-vectorized Cu2+. a and b: Enhancement of Cellular ROS level by ND-vectorized Cu2+. a: ROS generation, b: NAC protection (*p < 0.05, **p < 0.01, one-way ANOVA for comparison). c: TEM images of a typical L929 cell after incubation with NDs, Cu2+ and NDs-Cu2+ mixture for 24 h. Arrows indicate NDs (Scale bars = 2 μm). d: Flow cytometric analysis of L929 cells after incubation with NDs, Cu2+ and NDs-Cu2+ mixture for 24 h.

Mentions: Reactive oxygen species (ROS) associated oxidative stress is the general pathway for Cu2+ to induce bioeffects [37,38]. We then determined the intracellular ROS production to test whether this ND-Cu2+ interaction mediated cellular responses occurred through ROS accumulation induced by excessive Cu2+. Cells were loaded with the ROS measuring probe dichlorofluorescin diacetate (DCFH-DA). Results showed that after 24 h incubation, 25 μg/mL Cu2+ slightly increased the intracellular ROS production compared with that of control (~2.9 times) and at the other two time points (2 and 6 h), no significant increase in the intracellular ROS production was observed. However, when it is vectorized by NDs dramatically induced the production of ROS in time dependent manner, and the highest amount of ROS was generated after 24 h exposure, with ~25 fold increase over control (Figure 5a). Furthermore, pretreatment with N-Acetylcysteine (NAC, ROS scavenger [39]) for 2 h provided effective protection against Cu2+ induced ROS generation (Figure 5b).Figure 5


Nanodiamonds act as Trojan horse for intracellular delivery of metal ions to trigger cytotoxicity.

Zhu Y, Zhang Y, Shi G, Yang J, Zhang J, Li W, Li A, Tai R, Fang H, Fan C, Huang Q - Part Fibre Toxicol (2015)

Enhancement of bioeffects by ND-vectorized Cu2+. a and b: Enhancement of Cellular ROS level by ND-vectorized Cu2+. a: ROS generation, b: NAC protection (*p < 0.05, **p < 0.01, one-way ANOVA for comparison). c: TEM images of a typical L929 cell after incubation with NDs, Cu2+ and NDs-Cu2+ mixture for 24 h. Arrows indicate NDs (Scale bars = 2 μm). d: Flow cytometric analysis of L929 cells after incubation with NDs, Cu2+ and NDs-Cu2+ mixture for 24 h.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4374301&req=5

Fig5: Enhancement of bioeffects by ND-vectorized Cu2+. a and b: Enhancement of Cellular ROS level by ND-vectorized Cu2+. a: ROS generation, b: NAC protection (*p < 0.05, **p < 0.01, one-way ANOVA for comparison). c: TEM images of a typical L929 cell after incubation with NDs, Cu2+ and NDs-Cu2+ mixture for 24 h. Arrows indicate NDs (Scale bars = 2 μm). d: Flow cytometric analysis of L929 cells after incubation with NDs, Cu2+ and NDs-Cu2+ mixture for 24 h.
Mentions: Reactive oxygen species (ROS) associated oxidative stress is the general pathway for Cu2+ to induce bioeffects [37,38]. We then determined the intracellular ROS production to test whether this ND-Cu2+ interaction mediated cellular responses occurred through ROS accumulation induced by excessive Cu2+. Cells were loaded with the ROS measuring probe dichlorofluorescin diacetate (DCFH-DA). Results showed that after 24 h incubation, 25 μg/mL Cu2+ slightly increased the intracellular ROS production compared with that of control (~2.9 times) and at the other two time points (2 and 6 h), no significant increase in the intracellular ROS production was observed. However, when it is vectorized by NDs dramatically induced the production of ROS in time dependent manner, and the highest amount of ROS was generated after 24 h exposure, with ~25 fold increase over control (Figure 5a). Furthermore, pretreatment with N-Acetylcysteine (NAC, ROS scavenger [39]) for 2 h provided effective protection against Cu2+ induced ROS generation (Figure 5b).Figure 5

Bottom Line: In addition, theoretical calculation and molecular dynamics (MD) computation were used to illustrate the adsorption properties of different metal ion on NDs as well as release profile of ion from ND-ion complexes at different pH values.Detailed investigation of ND-Cu2+ interaction showed that the amount of released Cu2+ from ND-Cu2+ complexes at acidic lysosomal conditions was much higher than that at neutral conditions, leading to the elevation of intracellular ROS level, which triggered cytotoxicity.The present experimental and theoretical results provide useful insight into understanding of cytotoxicity triggered by nanoparticle-ion interactions, and open new ways in the interpretation of nanotoxicity.

View Article: PubMed Central - PubMed

Affiliation: Division of Physical Biology, and Bioimaging Center, Shanghai Synchrotron Radiation Facility, Shanghai Institute of Applied Physics, Chinese Academy of Sciences, Shanghai, 201800, China. zhuying@sinap.ac.cn.

ABSTRACT

Background: Nanomaterials hold great promise for applications in the delivery of various molecules with poor cell penetration, yet its potential for delivery of metal ions is rarely considered. Particularly, there is limited insight about the cytotoxicity triggered by nanoparticle-ion interactions. Oxidative stress is one of the major toxicological mechanisms for nanomaterials, and we propose that it may also contribute to nanoparticle-ion complexes induced cytotoxicity.

Methods: To explore the potential of nanodiamonds (NDs) as vehicles for metal ion delivery, we used a broad range of experimental techniques that aimed at getting a comprehensive assessment of cell responses after exposure of NDs, metal ions, or ND-ion mixture: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, Trypan blue exclusion text, optical microscope observation, synchrotron-based scanning transmission X-ray microscopy (STXM) and micro X-ray fluorescence (μXRF) microscopy, inductively coupled plasma-mass spectrometry (ICP-MS), reactive oxygen species (ROS) assay and transmission electron microscopy (TEM) observation. In addition, theoretical calculation and molecular dynamics (MD) computation were used to illustrate the adsorption properties of different metal ion on NDs as well as release profile of ion from ND-ion complexes at different pH values.

Results: The adsorption capacity of NDs for different metal ions was different, and the adsorption for Cu2+ was the most strong among divalent metal ions. These different ND-ion complexes then had different cytotoxicity by influencing the subsequent cellular responses. Detailed investigation of ND-Cu2+ interaction showed that the amount of released Cu2+ from ND-Cu2+ complexes at acidic lysosomal conditions was much higher than that at neutral conditions, leading to the elevation of intracellular ROS level, which triggered cytotoxicity. By theoretical approaches, we demonstrated that the functional carbon surface and cluster structures of NDs made them good vehicles for metal ions delivery.

Conclusions: NDs played the Trojan horse role by allowing large amounts of metal ions accumulate into living cells followed by subsequent release of ions in the interior of cells, which then led to cytotoxicity. The present experimental and theoretical results provide useful insight into understanding of cytotoxicity triggered by nanoparticle-ion interactions, and open new ways in the interpretation of nanotoxicity.

No MeSH data available.


Related in: MedlinePlus