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Comparative assessment of the apoptotic potential of silver nanoparticles synthesized by Bacillus tequilensis and Calocybe indica in MDA-MB-231 human breast cancer cells: targeting p53 for anticancer therapy.

Gurunathan S, Park JH, Han JW, Kim JH - Int J Nanomedicine (2015)

Bottom Line: This is especially true in the area of nanomedicine, due to physicochemical properties, such as mechanical, chemical, magnetic, optical, and electrical properties, compared with bulk materials.The first goal of this study was to produce silver nanoparticles (AgNPs) using two different biological resources as reducing agents, Bacillus tequilensis and Calocybe indica.Cells pretreated with pifithrin-alpha were protected from p53-mediated AgNPs-induced toxicity.

View Article: PubMed Central - PubMed

Affiliation: Department of Animal Biotechnology, Konkuk University, Seoul, Republic of Korea.

ABSTRACT

Background: Recently, the use of nanotechnology has been expanding very rapidly in diverse areas of research, such as consumer products, energy, materials, and medicine. This is especially true in the area of nanomedicine, due to physicochemical properties, such as mechanical, chemical, magnetic, optical, and electrical properties, compared with bulk materials. The first goal of this study was to produce silver nanoparticles (AgNPs) using two different biological resources as reducing agents, Bacillus tequilensis and Calocybe indica. The second goal was to investigate the apoptotic potential of the as-prepared AgNPs in breast cancer cells. The final goal was to investigate the role of p53 in the cellular response elicited by AgNPs.

Methods: The synthesis and characterization of AgNPs were assessed by various analytical techniques, including ultraviolet-visible (UV-vis) spectroscopy, X-ray diffraction (XRD), Fourier transform infrared (FTIR) spectroscopy, dynamic light scattering (DLS), and transmission electron microscopy (TEM). The apoptotic efficiency of AgNPs was confirmed using a series of assays, including cell viability, leakage of lactate dehydrogenase (LDH), production of reactive oxygen species (ROS), DNA fragmentation, mitochondrial membrane potential, and Western blot.

Results: The absorption spectrum of the yellow AgNPs showed the presence of nanoparticles. XRD and FTIR spectroscopy results confirmed the crystal structure and biomolecules involved in the synthesis of AgNPs. The AgNPs derived from bacteria and fungi showed distinguishable shapes, with an average size of 20 nm. Cell viability assays suggested a dose-dependent toxic effect of AgNPs, which was confirmed by leakage of LDH, activation of ROS, and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)-positive cells in MDA-MB-231 breast cancer cells. Western blot analyses revealed that AgNPs induce cellular apoptosis via activation of p53, p-Erk1/2, and caspase-3 signaling, and downregulation of Bcl-2. Cells pretreated with pifithrin-alpha were protected from p53-mediated AgNPs-induced toxicity.

Conclusion: We have demonstrated a simple approach for the synthesis of AgNPs using the novel strains B. tequilensis and C. indica, as well as their mechanism of cell death in a p53-dependent manner in MDA-MB-231 human breast cancer cells. The present findings could provide insight for the future development of a suitable anticancer drug, which may lead to the development of novel nanotherapeutic molecules for the treatment of cancers.

No MeSH data available.


Related in: MedlinePlus

B-AgNPs and F-AgNPs promote apoptosis.Notes: MDA-MB-231 cells were treated with respective IC50 concentrations of B-AgNPs or F-AgNPs for 24 hours. Fluorescent staining of cells was recorded. Representative images are shown for apoptotic DNA fragmentation (red staining) and corresponding nuclei (blue staining).Abbreviations: B-AgNPs, bacterium-derived AgNPs; DAPI, 4′,6-diamidino-2-phenylindole; F-AgNPs, fungus-derived AgNPs; IC50, half-maximal inhibitory concentration; TUNEL, terminal deoxynucleotidyl transferase dUTP nick end labeling.
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f9-ijn-10-4203: B-AgNPs and F-AgNPs promote apoptosis.Notes: MDA-MB-231 cells were treated with respective IC50 concentrations of B-AgNPs or F-AgNPs for 24 hours. Fluorescent staining of cells was recorded. Representative images are shown for apoptotic DNA fragmentation (red staining) and corresponding nuclei (blue staining).Abbreviations: B-AgNPs, bacterium-derived AgNPs; DAPI, 4′,6-diamidino-2-phenylindole; F-AgNPs, fungus-derived AgNPs; IC50, half-maximal inhibitory concentration; TUNEL, terminal deoxynucleotidyl transferase dUTP nick end labeling.

Mentions: ROS can act as signal molecules promoting cell-cycle progression and can induce oxidative DNA damage. DNA fragmentation is broadly considered a characteristic feature of apoptosis.81 Induction of apoptosis can be confirmed by two factors: irregular reduction in cell size, in which the cells are reduced and shrunken, and DNA fragmentation.9,65 Foldbjerg et al reported that AgNPs could elicit many biochemical and molecular changes in cultured cells.66 For example, AgNPs-induced DNA breakage was detected in cell lines by using a TUNEL assay. To confirm the apoptotic features induced by AgNPs in MDA-MB-231 cells, apoptotic DNA fragmentation was evaluated by fluorescence microscopy using the TUNEL assay (Figure 9). Treatment of MDA-MB-231 cells with B-AgNPs (10 μg/mL) and F-AgNPs (2 μg/mL) revealed a significant appearance of positively labeled cells, representing apoptotic DNA fragmentation. In control cultures, fewer or no apoptotic cells were observed. In contrast, DOX-treated cells showed a significant number of TUNEL-positive cells. Sriram et al demonstrated that Dalton’s lymphoma ascites cell lines treated with AgNPs exhibit DNA fragmentation.9 The toxicity of starch-coated AgNPs was studied using normal human lung fibroblast cells (IMR-90) and human glioblastoma cells (U251), and the results concluded that the toxicity of AgNPs influences cell morphology, cell viability, metabolic activity, and oxidative stress. Further, AgNPs reduced adenosine triphosphate (ATP) content of the cell, caused damage to mitochondria, and increased production of ROS in a dose-dependent manner.20 AgNPs can generate oxidative stress and lead to a series of cellular events, including reduced levels of glutathione (GSH), elevated lipid peroxidation, inflammation, DNA damage, altered cell cycle and proliferation capacity, and apoptosis and necrosis in various cell culture models.82


Comparative assessment of the apoptotic potential of silver nanoparticles synthesized by Bacillus tequilensis and Calocybe indica in MDA-MB-231 human breast cancer cells: targeting p53 for anticancer therapy.

Gurunathan S, Park JH, Han JW, Kim JH - Int J Nanomedicine (2015)

B-AgNPs and F-AgNPs promote apoptosis.Notes: MDA-MB-231 cells were treated with respective IC50 concentrations of B-AgNPs or F-AgNPs for 24 hours. Fluorescent staining of cells was recorded. Representative images are shown for apoptotic DNA fragmentation (red staining) and corresponding nuclei (blue staining).Abbreviations: B-AgNPs, bacterium-derived AgNPs; DAPI, 4′,6-diamidino-2-phenylindole; F-AgNPs, fungus-derived AgNPs; IC50, half-maximal inhibitory concentration; TUNEL, terminal deoxynucleotidyl transferase dUTP nick end labeling.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4494182&req=5

f9-ijn-10-4203: B-AgNPs and F-AgNPs promote apoptosis.Notes: MDA-MB-231 cells were treated with respective IC50 concentrations of B-AgNPs or F-AgNPs for 24 hours. Fluorescent staining of cells was recorded. Representative images are shown for apoptotic DNA fragmentation (red staining) and corresponding nuclei (blue staining).Abbreviations: B-AgNPs, bacterium-derived AgNPs; DAPI, 4′,6-diamidino-2-phenylindole; F-AgNPs, fungus-derived AgNPs; IC50, half-maximal inhibitory concentration; TUNEL, terminal deoxynucleotidyl transferase dUTP nick end labeling.
Mentions: ROS can act as signal molecules promoting cell-cycle progression and can induce oxidative DNA damage. DNA fragmentation is broadly considered a characteristic feature of apoptosis.81 Induction of apoptosis can be confirmed by two factors: irregular reduction in cell size, in which the cells are reduced and shrunken, and DNA fragmentation.9,65 Foldbjerg et al reported that AgNPs could elicit many biochemical and molecular changes in cultured cells.66 For example, AgNPs-induced DNA breakage was detected in cell lines by using a TUNEL assay. To confirm the apoptotic features induced by AgNPs in MDA-MB-231 cells, apoptotic DNA fragmentation was evaluated by fluorescence microscopy using the TUNEL assay (Figure 9). Treatment of MDA-MB-231 cells with B-AgNPs (10 μg/mL) and F-AgNPs (2 μg/mL) revealed a significant appearance of positively labeled cells, representing apoptotic DNA fragmentation. In control cultures, fewer or no apoptotic cells were observed. In contrast, DOX-treated cells showed a significant number of TUNEL-positive cells. Sriram et al demonstrated that Dalton’s lymphoma ascites cell lines treated with AgNPs exhibit DNA fragmentation.9 The toxicity of starch-coated AgNPs was studied using normal human lung fibroblast cells (IMR-90) and human glioblastoma cells (U251), and the results concluded that the toxicity of AgNPs influences cell morphology, cell viability, metabolic activity, and oxidative stress. Further, AgNPs reduced adenosine triphosphate (ATP) content of the cell, caused damage to mitochondria, and increased production of ROS in a dose-dependent manner.20 AgNPs can generate oxidative stress and lead to a series of cellular events, including reduced levels of glutathione (GSH), elevated lipid peroxidation, inflammation, DNA damage, altered cell cycle and proliferation capacity, and apoptosis and necrosis in various cell culture models.82

Bottom Line: This is especially true in the area of nanomedicine, due to physicochemical properties, such as mechanical, chemical, magnetic, optical, and electrical properties, compared with bulk materials.The first goal of this study was to produce silver nanoparticles (AgNPs) using two different biological resources as reducing agents, Bacillus tequilensis and Calocybe indica.Cells pretreated with pifithrin-alpha were protected from p53-mediated AgNPs-induced toxicity.

View Article: PubMed Central - PubMed

Affiliation: Department of Animal Biotechnology, Konkuk University, Seoul, Republic of Korea.

ABSTRACT

Background: Recently, the use of nanotechnology has been expanding very rapidly in diverse areas of research, such as consumer products, energy, materials, and medicine. This is especially true in the area of nanomedicine, due to physicochemical properties, such as mechanical, chemical, magnetic, optical, and electrical properties, compared with bulk materials. The first goal of this study was to produce silver nanoparticles (AgNPs) using two different biological resources as reducing agents, Bacillus tequilensis and Calocybe indica. The second goal was to investigate the apoptotic potential of the as-prepared AgNPs in breast cancer cells. The final goal was to investigate the role of p53 in the cellular response elicited by AgNPs.

Methods: The synthesis and characterization of AgNPs were assessed by various analytical techniques, including ultraviolet-visible (UV-vis) spectroscopy, X-ray diffraction (XRD), Fourier transform infrared (FTIR) spectroscopy, dynamic light scattering (DLS), and transmission electron microscopy (TEM). The apoptotic efficiency of AgNPs was confirmed using a series of assays, including cell viability, leakage of lactate dehydrogenase (LDH), production of reactive oxygen species (ROS), DNA fragmentation, mitochondrial membrane potential, and Western blot.

Results: The absorption spectrum of the yellow AgNPs showed the presence of nanoparticles. XRD and FTIR spectroscopy results confirmed the crystal structure and biomolecules involved in the synthesis of AgNPs. The AgNPs derived from bacteria and fungi showed distinguishable shapes, with an average size of 20 nm. Cell viability assays suggested a dose-dependent toxic effect of AgNPs, which was confirmed by leakage of LDH, activation of ROS, and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)-positive cells in MDA-MB-231 breast cancer cells. Western blot analyses revealed that AgNPs induce cellular apoptosis via activation of p53, p-Erk1/2, and caspase-3 signaling, and downregulation of Bcl-2. Cells pretreated with pifithrin-alpha were protected from p53-mediated AgNPs-induced toxicity.

Conclusion: We have demonstrated a simple approach for the synthesis of AgNPs using the novel strains B. tequilensis and C. indica, as well as their mechanism of cell death in a p53-dependent manner in MDA-MB-231 human breast cancer cells. The present findings could provide insight for the future development of a suitable anticancer drug, which may lead to the development of novel nanotherapeutic molecules for the treatment of cancers.

No MeSH data available.


Related in: MedlinePlus