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Potential of colloidal or silver nanoparticles to reduce the growth of B16F10 melanoma tumors


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Previosuly, we reported the cytotoxic effect of colloidal silver (AgC) on MCF-7 breast cancer cell line. However, there is scarce information on its antitumor potential. The aim of this study was to evaluate the anti-tumoral activity of colloidal silver (AgC) or silver nanoparticles (AgNPs) in a B16F10 melanoma mice model. In vitro, B16F10 cells were treated with different concentrations of AgC or AgNPs and cell viability was evaluated by MTT method, both treatments had cytotoxic effects against B16F10 cell line. In vivo, B16F10 melanoma cells (5 × 10 5) were implanted in six weeks old C57BL/6 mice. About 8 days after cells injection, the subcutaneous treatments were started with AgC or AgNPs, tumor volume and tumor weight were evaluated and the difference of treated groups and control demonstrated that melanoma tumor growth was significantly decreased. Our results suggest that AgC or AgNPs could be useful as an antiproliferative drug, inducing an impairment of tumoral growth.
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Vol. 7(22), pp. 2745-2750, 28 May, 2013
DOI: 10.5897/AJMR12.1968
ISSN 1996-0808 ©2013 Academic Journals
African Journal of Microbiology Research
Full Length Research Paper
Potential of colloidal or silver nanoparticles to reduce
the growth of B16F10 melanoma tumors
Sierra-Rivera C. A., Franco-Molina M. A.*, Mendoza-Gamboa E., Zapata-Benavides P., Tamez-
Guerra R. S. and Rodríguez-Padilla C.
Laboratorio de Inmunología y Virología, Departamento de Microbiología e Inmunología, Facultad de Ciencias Biológicas
de la Universidad Autónoma de Nuevo León, San Nicolás de los Garza, N. L. México.
Accepted 26 March, 2013
Previosuly, we reported the cytotoxic effect of colloidal silver (AgC) on MCF-7 breast cancer cell line.
However, there is scarce information on its antitumor potential. The aim of this study was to evaluate
the anti-tumoral activity of colloidal silver (AgC) or silver nanoparticles (AgNPs) in a B16F10 melanoma
mice model. In vitro, B16F10 cells were treated with different concentrations of AgC or AgNPs and cell
viability was evaluated by MTT method, both treatments had cytotoxic effects against B16F10 cell line.
In vivo, B16F10 melanoma cells (5 × 10
) were implanted in six weeks old C57BL/6 mice. About 8 days
after cells injection, the subcutaneous treatments were started with AgC or AgNPs, tumor volume and
tumor weight were evaluated and the difference of treated groups and control demonstrated that
melanoma tumor growth was significantly decreased. Our results suggest that AgC or AgNPs could be
useful as an antiproliferative drug, inducing an impairment of tumoral growth.
Key words: Colloidal silver, silver nanoparticles, melanoma, cancer, tumor.
The recent increase in the incidence of malignant
melanoma urges the development of more specific and
effective therapies. Historically, silver has been a major
therapeutic agent in medicine, especially in infectious
disease, including surgical infections (Alexander, 2009).
Since 1990, there has been a resurgence on the use of
AgC as an alternative medicine because of increased
resistance of bacteria to antibiotics, and the continuing
search for novel and affordable antimicrobial agents.
Previously, we reported that AgC has antitumor activity
through induction of apoptosis in MCF-7 breast cancer
cell line (Franco-Molina et al., 2010) and other studies
with AgNPs, by Sriram et al. (2010) demonstrated the
efficacy of biolo-synthesized AgNPs as an antitumor
agent against Dalton’s lymphoma ascites cell lines in vitro
and in vivo and the capacity to affect cellular viability of
human colon cancer cells (HT 29) (Sanpui et al., 2011).
Despite the fact that nanoparticles and colloidal particles
possess great potential for future clinical application
therapeutics, and this application generated interest in
exploring other metals for potential anti-cancer properties
(Bhattacharyya et al., 2011), there is scarce information
on AgC or AgNPs antitumor potential. This study is
relevant due to their use within health remedies. The aim
of the present study was to determine the effects of AgC
or AgNPs on murine melanoma tumorigenesis under in
vitro and in vivo conditions.
*Corresponding author. E-mail: Tel: 01152-81-83-29-41-15. Fax: 01152- 81-83-52-42-12.
Abbreviations: AgC, Colloidal silver; AgNPs, silver nanoparticles; MTT, thiazoly blue tetrazolium bromide.
2746 Afr. J. Microbiol. Res.
Figure 1. Schematic representation of the treatment protocols. Tumor time-line. 5x10
B16F10 cells were
inoculated subcutaneously into C57BL/6 mice on day 0, later beginning on day 8 after tumor cell
inoculation, the treatments with AgC (28 mg/kg) or AgNPs (1000 mg/kg) were administrated around tumor
with daily subcutaneous dose, the mice were monitored until 21 days post-cell inoculation and were finally
sacrificed by cervical dislocation.
In vivo experiments were conducted in male C57BL/6 mice of six
weeks old (18 to 25 g), housed at 22 ± 2°C under a 12/12 h
light/dark cycle with free access to food and water until use. All
animal handling and experimental procedures were conducted with
prior approval of the ethics committee on animal use of the
Facultad de Ciencias Biológicas, Universidad Autónoma de Nuevo
AgC or AgNPs
The grenetine-stabilized AgC (120 nm) was purchased from
MICRODYN (Mexico, D.F.) as a 0.35% stock solution. It was filtered
and diluted to a concentration of 17.5 µg/mL with DMEM/F-12
supplemented with 10% FBS. The AgNPs (10 nm) was purchased
from (Nanostructured & Amorphous Materials Inc. Houston, Tx.
USA); 5 mg/mL stock solution was prepared in DMEM-F12 supple-
mented with 10% FBS, previously sonicated during 15 min for its
homogenization and sterilized by filtration (0.2 µm filter, Millipore,
USA) in vitro for assays. For In vivo studies, 28 mg/kg (AgC) or
1000 mg/kg (AgNPs) per mice were daily administered subcuta-
neously around the tumor.
Cell viability
Cells (5 × 10
cells/well) were plated on 96 flat-bottom well plates,
and incubated 24 h at 37°C in 5% CO
atmosphere. After incuba-
tion, culture medium was removed, and AgC diluted in the same
medium was added at concentrations ranging from 0 to 17.5 µg/mL
and AgNPs at concentrations ranging from 0 to 5 mg/mL. The
plates were then incubated for 24 h at 37°C, and 5% CO
sphere. Thereafter, the supernatant was removed and cells were
washed twice with DMEM/F-12 medium. Cell viability was deter-
mined by the MTT method. Quantification was obtained by the
absorbance reading at a wavelength of 570 nm and cellular viability
was expressed as percentage. Results were given as the mean ±
SD of three independent experiments.
Melanoma cells implant
All animal procedures were made in Laboratorio de Inmunología de
la Facultad de Ciencia Biológicas de la UANL, in accordance with
Facultad de Ciencias Biológicas of the UANL Ethics Committee.
Briefly, 5 × 10
B16F10 melanoma cells were injected subcuta-
neously into C57BL/6 mice. When tumors were palpable (around 8
days after implanted) mice were treated around tumor with subcuta-
neous daily dose of 28 mg/Kg of AgC or 1000 mg/Kg of AgNPs.
The mice were monitored until 21 days post-cell inocula-tion and
were sacrificed by cervical dislocation (Figure 1). The tumors were
surgically collected, and body weight, volume, tumor weight and
metastasis were determined (metastasis was deter-mined during
the necropsy by findings of tumor cells in muscle, peritoneal cavity,
bowel and liver). Tumor volume was recorded using a measuring
gauge (PRETUL, USA) using the formula: volume = length x
Statistical analysis
Data represent the mean ±SD of triplicates from three independent
experiments. Statistical differences were obtained using the analy-
sis of variance, and the Dunnett's tests (SPSS v. 17.0 program).
The results were considered statistically significant if the *p value
was <0.05.
Cytotoxic activity of AgC or AgNPs on B16F10
melanoma cell line
AgC induced dose-dependent cytotoxic effect (7 to 17.5
µg/mL) on B16F10 cells (Figure 2). AgNPs induced dose-
dependent cytotoxic effect (1.5 to 5 mg/mL) on B16F10
melanoma cells in 24 h of incubation (Figure 3).
Treatment of B16F10 melanoma tumor
The B16F10 melanoma model was used to demonstrate
the therapeutic value of AgC or AgNPs. After the tumor
appearance by the eight day, the group mice were daily
treated with 28 mg/kg of AgC or AgNPs 1000 mg/kg,
respectively, by subcutaneous route around tumor. The
mice were sacrificed at 21 days of treatment (Figure 1).
There was no difference regarding the body weight bet-
ween treatments (control (25.00 g ± 2.78), AgNPs (21.30
g ± 1.14) and AgC (25.81 g ± 0.39) (Table 1 and Figure
4). However, these treatments significantly (p<0.05)
decreased the tumor weight {(AgNPs (0.51 ± 0.22 g) and
AgC (0.85 ± 0.64 g)} when compared with the control
(4.97 g ± 0.31) (Table 1 and Figure 4); and the tumor
Sierra-Rivera et al. 2747
Figure 2. B16F10 cell viability treated with AgC. B16F10 cells (5 x 10
cells/well) were cultured into
96 well plates and incubated overnight. Thereafter, the plates were treated with AgC concentrations
ranging from 0 to 17.5 µg/mL, and incubated for 24 h at 37°C, and 5% CO
atmosphere. Thereafter, a
MTT assay was performed. The optical density was determined at 570 nm. Data represent means of
triplicate samples with ±SD indicated. *p<0.05 as compared with untreated cells.
Figure 3. B16F10 cell viability treated with AgNPs. B16F10 cells (5 x 10
cells/well) were cultured
into 96 well plates and incubated overnight. Thereafter, the plates were treated with AgNPs
concentrations ranging from 0 to 5 mg/mL, and incubated for 24 h at 37°C, and 5% CO
atmosphere. Thereafter, a MTT assay was performed. The optical density was determined at 570
nm. Data represent means of triplicate samples with ±SD indicated. *p<0.05 as compared with
untreated cells.
volume was significantly (p<0.05) decreased {(AgNPs
(1.21 mm
± 0.68) and AgC (1.30 mm
± 0.73)} when
compared with the control (11.01 mm
± 0.86) (Table 1
and Figure 4). Regarding to the necropsy findings; the
treatments of AgC or AgNPs prevented the metastasis
from muscle, peritoneal cavity, bowel, and liver when
compared with control (Table 2). None mice showed
tumoral erradication in both groups treated. The mice
2748 Afr. J. Microbiol. Res.
Figure 4. Tumor growth decrease in mice treated with AgC or AgNPs. C57BL/6 mice bearing B16F10 tumor,
were treated daily with AgNP´s or AgC at doses of 28 and 1000 mg/kg, respectively. A) Control, B) AgC and
C) AgNPs, these pictures are representative mice groups.
Table 1. Comparison of volume-weight of tumors in mice treated with AgC or AgNPs.
Body weight (g ± SD)
Tumor weight (g ± SD)
Tumor volume (mm
± SD)
C57BL/6 mice bearing B16F10 tumor were daily treated with AgNPs or AgC at doses of 1000 or 28 mg/kg, respectively. In
this study, the mice were sacrificed at 21 days. *p<0.05 as compared to the controls.
Table 2. Incidence of metastasis in mice treated with AgC or AgNPs
Peritoneal cavity
C57BL/6 mice bearing B16F10 tumor, were treated daily with AgNPs or AgC at doses of 1000 or 28
mg/kg, respectively. In this study, the mice were sacrificed by cervical dislocation at 21 days; and the
metastasis was determined during the necropsy in muscle, peritoneal cavity, bowel and liver. +:
Metastasis, - : without metastasis.
treated with AgC showed a skin induration like fibrosis in
the zone treated for five days, this condition was ob-
served until sacrifice; the mice treated with AgNPs were
not affected by this symptoms.
Skin cancer is the most common form of cancer in the
United States. More than 3.5 million skin cancers in over
two million people are diagnosed annually. Each year
there are more new cases of skin cancer than the com-
bined incidence of cancers of the breast, prostate, lung
and colon. An estimated 123,590 new cases of mela-
noma were diagnosed in the US in 2011: 53,360 noninva-
sive (in situ) and 70,230 invasive, with nearly 8,790
resulting in death ( Some
cytotoxic agents used for its treatment are costly and
known to induce several side effects such as myelosup-
pression, anemia and most importantly the generation of
cellular resistance. For this, it is important to find alterna-
tive therapies or drugs to overcome these drawbacks
(Kim et al., 2007). Since ancient times, the silver has
been used to treat numerous diseases, mainly used in
antimicrobial agents, in treating wounds, burns and
catherter related infections (AshaRani et al., 2009),
however; it was until only a few years ago that there was
boom in taking different silver particles due to the rate of
exposure increasing progresively over the years when
engineered nanomaterials were extensively used in a
variety of industries including medical applications
(AshaRani et al., 2012). The use of smaller particles have
a wider tissue distribution, penetrate further within the
skin and intestine, are internalised to a greater extent,
and have a larger toxic potency (Johnston et al., 2010).
So the study of different silver particles is vital to explore
their therapeutic potential. Previously, we reported the
antitumor activity of colloidal silver on MCF-7 human
breast cancer cells (Franco-Molina et al., 2010), and for
this reason we decided to continue with a tumor model In
vivo. In the present study, our In vitro results demon-
strated that AgC or AgNPs significantly decreased in a
dose-dependent manner (p<0.05), the growth of B16F10
melanoma cells. The effects of cytotoxicity are similar to
those shown by other studies (Hsin et al., 2008) and the
mechanism of cell death probably are due to decreased
mitochondrial membrane potential, inducing apoptotic
death (Franco-Molina et al., 2010). Although, the exact
mechanism of action of silver particles is unknown; some
reports showed that AgNP uptake occurs mainly through
endocytosis where clathrin mediated process and macro-
pinocytosis were involved, the nuclear deposition of
AgNP is unknown, but the AgNP treatment leads to
changes in the cell membrane permeability, facilitating
the entry of Ca
ions which activate enzymes like pro-
teases and endonuclease that increase toxicity, resulting
in mitochondrial membrane dysfunction and reactive
oxygen species production and oxidative stress, damage
to DNA can be induced through binding of DNA or via
oxidative to DNA, reducing DNA synthesis and producing
chromosomal aberrations, errors in chromosome segre-
gation and production of micronuclei, leading to cell death
(AshaRani et al., 2009), mainly observed in tumor cells
and not in normal cells (Franco-Molina et al., 2010). It is
important to notice that major doses were used to induce
cytotoxic effect by AgNPs (1.5 to 5 mg/mL) when com-
pared with AgC (7 to 17.5 µg/mL) on cancer cells. This
findings were probably due to the fact that AgNPs treated
cells have limited exposure to Ag ions because AgNPs
solution contained a minimun amount of free Ag ions.
However, it has been suggested that AgNPs and Ag can
induce cell death in vitro through a ROS production. It
has been demonstrated that there are some differences
in their mechanism of action, example is that Ag induced
metallothionein 1b (MT1b) and AgNPs did not. Ag also is
capable of inducing a major production of oxidative stress
Sierra-Rivera et al. 2749
related glutathione peroxidase and catalase expression
compared with AgNPs. This mechanism of action could
indicate that AgC appeared more toxic than AgNPs such
has been demonstrated by Hsin et al., 2008.
“In vivo we found that AgC or AgNPs have the poten-
tial to impair the growth tumor, since AgNPs is better than
AgC treatment in reducing the tumoral volume and
weight. These finding could be correlated with another
study where the AgNPs had the potential of inhibiting the
VEGF in a model employing bovine retinal endothelial
cells in vitro and angiogenesis in a mice model in vivo
(Gurunathan et al., 2009). At the beginning of the study,
we observed that intraperitoneal injection of AgC (28
mg/kg) induced mice exhibited nervous symptoms after
15 s of administration, like jumping and disnea by 10 min
and lethargy during all day long (data not shown), by this
reason the use of subcutaneous administration at doses
of 28 mg/kg, without side effects was decided, avoiding
increases of the doses like that used with AgNPs. There
are reports that show adverse effects when doses higher
than 100 mg/kg of AgC were used (Faust, 1992). And the
adverse effects could be possible because the Ag can
enter through the blood-brain barrier and accumulate in
large motoneurones in the brain stem and spinal cord,
neurons in cerebellar nuclei and glia; and the toxic symp-
toms such as cerebral ataxia are associated with pro-
longed exposure to silver (Panyala et al., 2008), which is
different from the findings previously mentioned by us.
In this experiment, we used the highest doses reported
for AgNPs because in a study by Rahman et al. (2009)
on the effects of AgNPs (25 nm) on gene expression in
different regions of the mouse brain where the particles
were administered to adult male mice route intraperito-
neal injection at doses of 100, 500, or 1000 mg/kg for 24
h, array data indicated changes in the expression of
genes in the caudate nucleus, frontal cortex and hippo-
campus of mice treated with the AgNPs. Analysis of
these changes led the authors to suggest that AgNPs
may produce neurotoxicity by stimulating oxidative stress
generation and altered gene expression, leading to apop-
tosis (Rahman et al., 2009). In the present study, despite
the use of AgNPs daily for thirteen days, we did not find
adverse effects, like nervous symptoms associated at its
Although, the mechanism of the antitumoral action of
AgC or AgNPs is not properly understood, it has been
reported that heavy metals react with proteins by getting
attached to the thiol group and the proteins get inacti-
vated through a mechanism implicated in avoiding the
cellular proliferation of cancer cells (Liau et al., 1997).
With these results, we can confirm the potential of AgC
or AgNPs on this melanoma tumor model suggesting
them as a potential agent for use in cancer treatment.
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... AgNP have many biomedical applications, including acting as carriers for antitumor agents [23] and mediating photodynamic therapy [24]. The ability of AgNP to directly kill melanoma cells was first reported in 2013 and studies have suggested that AgNP can induce melanoma cell death through apoptosis in a dose-dependent manner in vitro [25,26]. A dose-dependent effect has also been shown in vivo where AgNP were administered subcutaneously to melanoma-challenged mice; and high concentrations of AgNP (12 mg/kg) were also shown to be safe [25]. ...
... The use of AgNP as an anti-melanoma agent was first reported in 2013 [26]. However, the physicochemical properties of AgNP can vary depending on size, shape, and capping agent used [20,38,39]. ...
... Melanoma skin cancer is responsible for the large majority of deaths related to skin cancer, with approximately 10,000 people expected to die from melanoma every year in the United States [53]. One of the most common treatments of melanoma cancer involves cell transfer therapies, based on antitumor lymphocytes and cytotoxic agents [54,55]. The cytotoxic agents might cause, for example, anemia and generation of cellular resistance [55]. ...
... One of the most common treatments of melanoma cancer involves cell transfer therapies, based on antitumor lymphocytes and cytotoxic agents [54,55]. The cytotoxic agents might cause, for example, anemia and generation of cellular resistance [55]. A remedy to these drawbacks may be the use of AgNPs in the treatment of cancer. ...
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An innovative and environmental friendly method for the synthesis of size-controlled silver nanoparticles (AgNPs) is presented. Pectin (PEC) stabilized AgNPs are synthesized in a reaction discharge system where pm-rf-APGD (pulse modulated radio frequency atmospheric pressure glow discharge) is operated in contact with flowing liquid electrode. AgNPs synthesized under defined operating conditions exhibit an average size of 41.62±12.08 nm and 10.38±4.56 nm, as determined by Dynamic Light Scattering (DLS) and Transmission Electron Microscopy (TEM), respectively. Energy-dispersive X-ray spectroscopy (EDS) confirms that the NPs are composed of metallic Ag. Furthermore, the ξ-potential of the AgNPs is shown to be -43.11±0.96 mV, which will facilitate their application in biological systems. Between 70% and 90% of the cancerous of the human melanoma Hs 294T cell line undergo necrosis following treatment with the synthesized AgNPs. Furthermore, optical emission spectrometry identifies reactive species, like NO, NH, N2, O, H, as pm-rf-APGD produced compounds that may be involved in the reduction of the Ag(I) ions.
... The cytotoxic effect of colloidal silver (Ag), sodium dichloroacetate (DCA), and their combination was evaluated against B16F10 murine melanoma cells. Our results show that Ag has antiproliferative effects against B16F10 cells, as previously reported by our research group [13]. Further reports of the cytotoxic activity of silver against melanoma cells refer to silver nanoparticles, although the proposed toxicity mechanism remains the same [14]. ...
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Our main focus was to evaluate the efficacy of silver and sodium dichloroacetate as dual-function agents to be used in melanoma treatment. This strategy is designed to increase the activity of these two compounds that affect DNA integrity and the mitochondria at different levels. Furthermore, we evaluated if the cell death mechanism induced by our treatments was immunogenic cell death. To evaluate antitumor efficacy, we assessed tumor volume and production of tumor necrosis factor-α, nuclear factor κ B (both by ELISA), and nitric oxide levels (Nitrate/Nitrite colorimetric assay kit); for immunogenic cell death, we evaluated the release of danger-associated molecular patterns using immunohistochemistry and flow cytometry, as well as an in vivo challenge. Our results showed that the combination of colloidal silver and sodium dichloroacetate is more effective than each treatment alone and that the antitumor mechanism is not through immunogenic cell death. Furthermore, this study can broadly contribute to the development of dichloroacetate-loaded silver nanoparticles and to the design targeted pharmacological formulations to fight melanoma as well as other types of cancer.
... TAT-AgNPs of 8 nm with an IC 50 value 52 nM increases the antitumor activity by 24-fold. Similarly, Sierra-Rivera et al. (2013) demonstrated the antitumor potential of grenetinestabilized colloidal silver of 120 nm and AgNPs of size 10 nm on B16F10 melanoma tumor. Dose-dependent cytotoxicity was reported at different concentrations for both forms of silver. ...
... The generated ROS and AgNPs escape from the lysosomes and then they disrupt the mitochondria. This event leads to the generation of more ROS, ultimately leading to DNA damage and cell death [12][13][14][15][16][17] . AgNPs were reported to possess potent anti-angiogenic effects via inhibition of vascular endothelial growth factor (VEGF)-induced angiogenesis both in-vitro and in-vivo 18 . ...
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The current study aimed at preparing AgNPs and three different core-shell silver/polymeric NPs composed of Ag core and three different polymeric shells: polyvinyl alcohol (PVA), polyethylene glycol (PEG) and polyvinylpyrrolidone (PVP). Thereafter, the core/shell NPs were loaded with a chemotherapeutic agent doxorubicin (DOX). Finally, the cytotoxic effects of the different core-shell Ag/polymeric NPs-based combinatorial therapeutics were tested in-vitro against breast cancer (MCF-7) and human fibroblast (1BR hTERT) cell lines. AgNPs, Ag/PVA and Ag/PVP NPs were more cytotoxic to MCF-7 cells than normal fibroblasts, as well as DOX-Ag, DOX-Ag/PVA, DOX-Ag/PEG and DOX-Ag/PVP nanocarriers (NCs). Notably, low dosage of core-shell DOX-loaded Ag/polymeric nanocarriers (NCs) exhibited a synergic anticancer activity, with DOX-Ag/PVP being the most cytotoxic. We believe that the prepared NPs-based combinatorial therapy showed a significant enhanced cytotoxic effect against breast cancer cells. Future studies on NPs-based combinatorial therapy may aid in formulating a novel and more effective cancer therapeutics.
... TAT-AgNPs of 8 nm with an IC 50 value 52 nM increases the antitumor activity by 24-fold. Similarly, Sierra-Rivera et al. (2013) demonstrated the antitumor potential of grenetinestabilized colloidal silver of 120 nm and AgNPs of size 10 nm on B16F10 melanoma tumor. Dose-dependent cytotoxicity was reported at different concentrations for both forms of silver. ...
Full-text available
Noble metals and their compounds have been used as therapeutic agents from the ancient time in medicine for the treatment of various infections. Recently, much progress has been made in the field of nanobiotechnology towards the development of different kinds of nanomaterials with a wide range of applications. Among the metal nanoparticles, noble metal nanoparticles have demonstrated potential biomedical applications. Due to the small size, nanoparticles can easily interact with biomolecules both at surface and inside cells, yielding better signals and target specificity for diagnostics and therapeutics. Noble metal nanoparticles inspired the researchers due to their remarkable role in detection and treatment of dreadful diseases. In this review, we have attempted to focus on the biomedical applications of noble metal nanoparticles particularly, silver, gold, and platinum in diagnosis and treatment of dreaded diseases such as cancer, human immunodeficiency virus (HIV), tuberculosis (TB), and Parkinson disease. In addition, the role of silver nanoparticles (AgNPs) such as novel antimicrobials, gold nanoparticles (AuNPs) such as efficient drug carrier, uses of platinum nanoparticles (PtNPs) in bone allograft, dentistry, etc. have been critically reviewed. Moreover, the toxicity due to the use of metal nanoparticles and some unsolved challenges in the field have been discussed with their possible solutions.
The aim of this report was to investigate the antitumor and apoptotic effects of silver nanoparticles (AgNPs) on the Dalton's ascites lymphoma cells in vivo. Thirty Swiss albino male mice were assigned into five groups of six each. Group I were intact animals. Group II animals served as tumor control injected with DAL cells intraperitonially. Group III induced animals received plant extract (17mg/kg BW) and Group IV induced animals received AgNPs (35μg/kg BW). Group V induced animals received standard anticancer drug 5-Fluorouracil (5-FU, 20μg/kg BW). The treatment period was 10 days excluding the day of tumor injection. Tumor cells were collected after euthanizing the animals and real-time PCR was used to analyze p53, caspase-3, 8, 9, 12 and cytochrome C expressions. Results indicate that the AgNPs were efficient in prolongation of life span, reduction of tumor volume and body weight in tumor animals. All the apoptotic genes were upregulated by treatment with AgNPs. To conclude, the present study elicits that AgNPs are potent in antitumor activity and the molecular mechanism is by the induction of apoptosis through the mitochondrial dependent and independent pathways.
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Investigating the cellular and molecular signatures in eukaryotic cells following exposure to nanoparticles will further our understanding on the mechanisms mediating nanoparticle induced effects. This study illustrates the molecular effects of silver nanoparticles (Ag-np) in normal human lung cells, IMR-90 and human brain cancer cells, U251 with emphasis on gene expression, induction of inflammatory mediators and the interaction of Ag-np with cytosolic proteins. We report that silver nanoparticles are capable of adsorbing cytosolic proteins on their surface that may influence the function of intracellular factors. Gene and protein expression profiles of Ag-np exposed cells revealed up regulation of many DNA damage response genes such as Gadd 45 in both the cell types and ATR in cancer cells. Moreover, down regulation of genes necessary for cell cycle progression (cyclin B and cyclin E) and DNA damage response/repair (XRCC1 and 3, FEN1, RAD51C, RPA1) was observed in both the cell lines. Double strand DNA damage was observed in a dose dependant manner as evidenced in γH2AX foci assay. There was a down regulation of p53 and PCNA in treated cells. Cancer cells in particular showed a concentration dependant increase in phosphorylated p53 accompanied by the cleavage of caspase 3 and PARP. Our results demonstrate the involvement of NFκB and MAP kinase pathway in response to Ag-np exposure. Up regulation of pro-inflammatory cytokines such as interleukins (IL-8, IL-6), macrophage colony stimulating factor, macrophage inflammatory protein in fibroblasts following Ag-np exposure were also observed. In summary, Ag-np can modulate gene expression and protein functions in IMR-90 cells and U251 cells, leading to defective DNA repair, proliferation arrest and inflammatory response. The observed changes could also be due to its capability to adsorb cytosolic proteins on its surface.
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Nanotechnology is an evolving field with enormous potential for biomedical applications. The growing interest to use inorganic nanoparticles in medicine is due to the unique size- and shape-dependent optoelectronic properties. Herein, we will focus on gold, silver and platinum nanoparticles, discussing recent developments for therapeutic applications with regard to cancer in terms of nanoparticles being used as a delivery vehicle as well as therapeutic agents. We will also discuss some of the key challenges to be addressed in future studies.
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Colloidal silver has been used as an antimicrobial and disinfectant agent. However, there is scarce information on its antitumor potential. The aim of this study was to determine if colloidal silver had cytotoxic effects on MCF-7 breast cancer cells and its mechanism of cell death. MCF-7 breast cancer cells were treated with colloidal silver (ranged from 1.75 to 17.5 ng/mL) for 5 h at 37°C and 5% CO2 atmosphere. Cell Viability was evaluated by trypan blue exclusion method and the mechanism of cell death through detection of mono-oligonucleosomes using an ELISA kit and TUNEL assay. The production of NO, LDH, and Gpx, SOD, CAT, and Total antioxidant activities were evaluated by colorimetric assays. Colloidal silver had dose-dependent cytotoxic effect in MCF-7 breast cancer cells through induction of apoptosis, shown an LD50 (3.5 ng/mL) and LD100 (14 ng/mL) (*P < 0.05), significantly decreased LDH (*P < 0.05) and significantly increased SOD (*P < 0.05) activities. However, the NO production, and Gpx, CAT, and Total antioxidant activities were not affected in MCF-7 breast cancer cells. PBMC were not altered by colloidal silver. The present results showed that colloidal silver might be a potential alternative agent for human breast cancer therapy.
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Nanomedicine concerns the use of precision-engineered nanomaterials to develop novel therapeutic and diagnostic modalities for human use. The present study demonstrates the efficacy of biologically synthesized silver nanoparticles (AgNPs) as an antitumor agent using Dalton's lymphoma ascites (DLA) cell lines in vitro and in vivo. The AgNPs showed dose- dependent cytotoxicity against DLA cells through activation of the caspase 3 enzyme, leading to induction of apoptosis which was further confirmed through resulting nuclear fragmentation. Acute toxicity, ie, convulsions, hyperactivity and chronic toxicity such as increased body weight and abnormal hematologic parameters did not occur. AgNPs significantly increased the survival time in the tumor mouse model by about 50% in comparison with tumor controls. AgNPs also decreased the volume of ascitic fluid in tumor-bearing mice by 65%, thereby returning body weight to normal. Elevated white blood cell and platelet counts in ascitic fluid from the tumor-bearing mice were brought to near-normal range. Histopathologic analysis of ascitic fluid showed a reduction in DLA cell count in tumor-bearing mice treated with AgNPs. These findings confirm the antitumor properties of AgNPs, and suggest that they may be a cost-effective alternative in the treatment of cancer and angiogenesis-related disorders.
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This review is concerned with evaluating the toxicity associated with human exposure to silver and gold nanoparticles (NPs), due to the relative abundance of toxicity data available for these particles, when compared to other metal particulates. This has allowed knowledge on the current understanding of the field to be gained, and has demonstrated where gaps in knowledge are. It is anticipated that evaluating the hazards associated with silver and gold particles will ultimately enable risk assessments to be completed, by combining this information with knowledge on the level of human exposure. The quantity of available hazard information for metals is greatest for silver particulates, due to its widespread inclusion within a number of diverse products (including clothes and wound dressings), which primarily arises from its antibacterial behaviour. Gold has been used on numerous occasions to assess the biodistribution and cellular uptake of NPs following exposure. Inflammatory, oxidative, genotoxic, and cytotoxic consequences are associated with silver particulate exposure, and are inherently linked. The primary site of gold and silver particulate accumulation has been consistently demonstrated to be the liver, and it is therefore relevant that a number of in vitro investigations have focused on this potential target organ. However, in general there is a lack of in vivo and in vitro toxicity information that allows correlations between the findings to be made. Instead a focus on the tissue distribution of particles following exposure is evident within the available literature, which can be useful in directing appropriate in vitro experimentation by revealing potential target sites of toxicity. The experimental design has the potential to impact on the toxicological observations, and in particular the use of excessively high particle concentrations has been observed. As witnessed for other particle types, gold and silver particle sizes are influential in dictating the observed toxicity, with smaller particles exhibiting a greater response than their larger counterparts, and this is likely to be driven by differences in particle surface area, when administered at an equal-mass dose. A major obstacle, at present, is deciphering whether the responses related to silver nanoparticulate exposure derive from their small size, or particle dissolution contributes to the observed toxicity. Alternatively, a combination of both may be responsible, as the release of ions would be expected to be greater for smaller particles.
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Background Nanoparticles possess exceptional physical and chemical properties which led to rapid commercialisation. Silver nanoparticles (Ag-np) are among the most commercialised nanoparticles due to their antimicrobial potential. Ag-np based cosmetics, therapeutic agents and household products are in wide use, which raised a public concern regarding their safety associated with human and environmental use. No safety regulations are in practice for the use of these nanomaterials. The interactions of nanomaterials with cells, uptake mechanisms, distribution, excretion, toxicological endpoints and mechanism of action remain unanswered. Results Normal human lung fibroblasts (IMR-90) and human glioblastoma cells (U251) were exposed to different doses of Ag-nps in vitro. Uptake of Ag-nps occurred mainly through endocytosis (clathrin mediated process and macropinocytosis), accompanied by a time dependent increase in exocytosis rate. The electron micrographs revealed a uniform intracellular distribution of Ag-np both in cytoplasm and nucleus. Ag-np treated cells exhibited chromosome instability and mitotic arrest in human cells. There was efficient recovery from arrest in normal human fibroblasts whereas the cancer cells ceased to proliferate. Toxicity of Ag-np is mediated through intracellular calcium (Ca2+) transients along with significant alterations in cell morphology and spreading and surface ruffling. Down regulation of major actin binding protein, filamin was observed after Ag-np exposure. Ag-np induced stress resulted in the up regulation of metallothionein and heme oxygenase -1 genes. Conclusion Here, we demonstrate that uptake of Ag-np occurs mainly through clathrin mediated endocytosis and macropinocytosis. Our results suggest that cancer cells are susceptible to damage with lack of recovery from Ag-np-induced stress. Ag-np is found to be acting through intracellular calcium transients and chromosomal aberrations, either directly or through activation of catabolic enzymes. The signalling cascades are believed to play key roles in cytoskeleton deformations and ultimately to inhibit cell proliferation.
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Nanotechnology is a rapidly growing science of producing and utilizing nano-sized particles that measure in nanometers. These nanomaterials are already having an impact on health care. Now-a-days we are using nanoproducts in various fields. Of these, silver nanoparticles are playing a major role in the field of nanotechnology and nanomedicine. Their unique size-dependent properties make these materials superior and indispensable as they show unusual physical, chemical and biological properties. Silver nanoparticles have potential antimicrobial activity towards many pathogenic microbes. Along with this antimicrobial activity, silver nanoparticles are showing unacceptable toxic effects on human health and the environment. The chronic exposure to silver causes adverse effects such as permanent bluish-grey discoloration of the skin (argyria) and eyes (argyrosis). Besides argyria and argyrosis, exposure to soluble silver compounds may produce other toxic effects like liver and kidney damage, irritation of the eyes, skin, respiratory and intestinal tract and changes to blood cells. This review summarizes the hazardous effects of silver nanoparticles in the environment and theirs toxic effects on human health.
Nanoparticles are small scale substances (<100 nm) used in biomedical applications, electronics, and energy production. Increased exposure to nanoparticles being produced in large-scale industry facilities elicits concerns for the toxicity of certain classes of nanoparticles. This study evaluated the effects of silver-25 nm (Ag-25) nanoparticles on gene expression in different regions of the mouse brain. Adult-male C57BL/6N mice were administered (i.p.) 100 mg/kg, 500 mg/kg or 1000 mg/kg Ag-25 and sacrificed after 24 h. Regions from the brain were rapidly removed and dissected into caudate nucleus, frontal cortex and hippocampus. Total RNA was isolated from each of the three brain regions collected and real-time RT-PCR analysis was performed using Mouse Oxidative Stress and Antioxidant Defense Arrays. Array data revealed the expression of genes varied in the caudate nucleus, frontal cortex and hippocampus of mice when treated with Ag-25. The data suggest that Ag-25 nanoparticles may produce neurotoxicity by generating free radical-induced oxidative stress and by altering gene expression, producing apoptosis and neurotoxicity.
We report the development of a chitosan nanocarrier (NC)-based delivery of silver nanoparticles (Ag NPs) to mammalian cells for induction of apoptosis at very low concentrations of the NPs. The cytotoxic efficacy of the Ag NP-nanocarrier (Ag-CS NC) system in human colon cancer cells (HT 29) was examined by morphological analyses and biochemical assays. Cell viability assay demonstrated that the concentration of Ag NPs required to reduce the viability of HT 29 cells by 50% was 0.33 μg mL(-1), much less than in previously reported data. The efficient induction of apoptosis by Ag-CS NCs was confirmed by flow cytometry. Additionally, the characteristic nuclear and morphological changes during apoptotic cell death were investigated by fluorescence and scanning electron microscopy (SEM), respectively. The involvement of mitochondrial pathway of cell death in the Ag-CS NCs induced apoptosis was evident from the depolarization of mitochondrial membrane potential (ΔΨ(m)). Real time quantitative RT-PCR analysis demonstrated the up-regulation of caspase 3 expression which was further reflected in the formation of oligo-nucleosomal DNA "ladders" in Ag-CS NCs treated cells, indicating the important role of caspases in the present apoptotic process. The increased production of intracellular ROS due to Ag-CS NCs treatment indicated that the oxidative stress could augment the induction of apoptosis in HT 29 cells in addition to classical caspase signaling pathway. The use of significantly low concentration of Ag NPs impregnated in chitosan nanocarrier is a much superior approach in comparison to the use of free Ag NPs in cancer therapy.
Angiogenesis is an important phenomenon involved in normal growth and wound healing processes. An imbalance of the growth factors involved in this process, however, causes the acceleration of several diseases including malignant, ocular, and inflammatory diseases. Inhibiting angiogenesis through interfering in its pathway is a promising methodology to hinder the progression of these diseases. The function and mechanism of silver nanoparticles (Ag-NPs) in angiogenesis have not been elucidated to date. PEDF is suggested to be a potent anti-angiogenic agent. In this study, we postulated that Ag-NPs might have the ability to inhibit angiogenesis, the pivotal step in tumor growth, invasiveness, and metastasis. We have demonstrated that Ag-NPs could also inhibit vascular endothelial growth factor (VEGF) induced cell proliferation, migration, and capillary-like tube formation of bovine retinal endothelial cells like PEDF. In addition, Ag-NPs effectively inhibited the formation of new blood microvessels induced by VEGF in the mouse Matrigel plug assay. To understand the underlying mechanism of Ag-NPs on the inhibitory effect of angiogenesis, we showed that Ag-NPs could inhibit the activation of PI3K/Akt. Together, our results indicate that Ag-NPs can act as an anti-angiogenic molecule by targeting the activation of PI3K/Akt signaling pathways.