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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

PFT-α inhibits B-AgNPs- and F-AgNPs-induced cell death in a p53-dependent manner.Notes: Cells were pretreated with a p53 inhibitor (PFT-α, 10 μM) for 1 hour and then incubated with respective IC50 concentrations of B-AgNPs or F-AgNPs for 24 hours. Effects on cell viability (A) and protein expression of p-p53 (B) are shown as mean ± SD of three independent experiments. Lane 1 shows control; lane 2 shows B-AgNPs; lane 3 shows F-AgNPs; lane 4 shows B-AgNPs with PFT-α; and lane 5 shows F-AgNPs with PFT-α.Abbreviations: B-AgNPs, bacterium-derived AgNPs; Con, control; F-AgNPs, fungus-derived AgNPs; IC50, half-maximal inhibitory concentration; PFT-α, pifithrin-alpha; SD, standard deviation.
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f12-ijn-10-4203: PFT-α inhibits B-AgNPs- and F-AgNPs-induced cell death in a p53-dependent manner.Notes: Cells were pretreated with a p53 inhibitor (PFT-α, 10 μM) for 1 hour and then incubated with respective IC50 concentrations of B-AgNPs or F-AgNPs for 24 hours. Effects on cell viability (A) and protein expression of p-p53 (B) are shown as mean ± SD of three independent experiments. Lane 1 shows control; lane 2 shows B-AgNPs; lane 3 shows F-AgNPs; lane 4 shows B-AgNPs with PFT-α; and lane 5 shows F-AgNPs with PFT-α.Abbreviations: B-AgNPs, bacterium-derived AgNPs; Con, control; F-AgNPs, fungus-derived AgNPs; IC50, half-maximal inhibitory concentration; PFT-α, pifithrin-alpha; SD, standard deviation.

Mentions: The small-molecule PFT-α has been reported to inhibit p53 function and protect against a variety of genotoxic agents.95 PFT-α, an inhibitor of p53, is known to reversibly inhibit p53 transcriptional activity and has been used for the characterization of p53 function in various experimental systems.96–99 Culmsee et al showed that DU145 cells treated with PFT-α exhibited reduced resveratrol-induced activation of p53, decreased p53 binding to DNA, and decreased levels of Bax, a target gene for p53.100 In this manuscript, previous experiments provided evidence that B-AgNPs- and F-AgNPs-induced cell death in MDA-MB-231 cells was mediated specifically by p53. To ascertain whether PFT-α suppressed p53-dependent cell death, we investigated whether PFT-α can rescue the apoptosis induced by AgNPs by pretreating cells with PFT-α and then measuring cell viability. The results clearly indicate that cells were rescued from B-AgNPs- and F-AgNPs-induced p53-mediated cell death when pretreated with PFT-α (Figure 12A). In addition, we performed Western blot analysis to confirm the inhibitory effect of PFT-α in the AgNPs-induced phosphorylation of p53. The results suggest that PFT-α inhibits the B-AgNPs-and F-AgNPs-induced phosphorylation of p53 (Figure 12B). Altogether, the results from the present study suggest that both B-AgNPs and F-AgNPs induce apoptosis through p53, which can be protected by PFT-α.


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)

PFT-α inhibits B-AgNPs- and F-AgNPs-induced cell death in a p53-dependent manner.Notes: Cells were pretreated with a p53 inhibitor (PFT-α, 10 μM) for 1 hour and then incubated with respective IC50 concentrations of B-AgNPs or F-AgNPs for 24 hours. Effects on cell viability (A) and protein expression of p-p53 (B) are shown as mean ± SD of three independent experiments. Lane 1 shows control; lane 2 shows B-AgNPs; lane 3 shows F-AgNPs; lane 4 shows B-AgNPs with PFT-α; and lane 5 shows F-AgNPs with PFT-α.Abbreviations: B-AgNPs, bacterium-derived AgNPs; Con, control; F-AgNPs, fungus-derived AgNPs; IC50, half-maximal inhibitory concentration; PFT-α, pifithrin-alpha; SD, standard deviation.
© Copyright Policy
Related In: Results  -  Collection

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

f12-ijn-10-4203: PFT-α inhibits B-AgNPs- and F-AgNPs-induced cell death in a p53-dependent manner.Notes: Cells were pretreated with a p53 inhibitor (PFT-α, 10 μM) for 1 hour and then incubated with respective IC50 concentrations of B-AgNPs or F-AgNPs for 24 hours. Effects on cell viability (A) and protein expression of p-p53 (B) are shown as mean ± SD of three independent experiments. Lane 1 shows control; lane 2 shows B-AgNPs; lane 3 shows F-AgNPs; lane 4 shows B-AgNPs with PFT-α; and lane 5 shows F-AgNPs with PFT-α.Abbreviations: B-AgNPs, bacterium-derived AgNPs; Con, control; F-AgNPs, fungus-derived AgNPs; IC50, half-maximal inhibitory concentration; PFT-α, pifithrin-alpha; SD, standard deviation.
Mentions: The small-molecule PFT-α has been reported to inhibit p53 function and protect against a variety of genotoxic agents.95 PFT-α, an inhibitor of p53, is known to reversibly inhibit p53 transcriptional activity and has been used for the characterization of p53 function in various experimental systems.96–99 Culmsee et al showed that DU145 cells treated with PFT-α exhibited reduced resveratrol-induced activation of p53, decreased p53 binding to DNA, and decreased levels of Bax, a target gene for p53.100 In this manuscript, previous experiments provided evidence that B-AgNPs- and F-AgNPs-induced cell death in MDA-MB-231 cells was mediated specifically by p53. To ascertain whether PFT-α suppressed p53-dependent cell death, we investigated whether PFT-α can rescue the apoptosis induced by AgNPs by pretreating cells with PFT-α and then measuring cell viability. The results clearly indicate that cells were rescued from B-AgNPs- and F-AgNPs-induced p53-mediated cell death when pretreated with PFT-α (Figure 12A). In addition, we performed Western blot analysis to confirm the inhibitory effect of PFT-α in the AgNPs-induced phosphorylation of p53. The results suggest that PFT-α inhibits the B-AgNPs-and F-AgNPs-induced phosphorylation of p53 (Figure 12B). Altogether, the results from the present study suggest that both B-AgNPs and F-AgNPs induce apoptosis through p53, which can be protected by PFT-α.

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