الفهرس 
Response of organisms to stressesOnly 14 pages are availabe for public view
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Abstract
This study was carried out at the Department of Genetics
(Drosophila Lab) and the Molecular Genetic Laboratory, Faculty of
Agriculture, Ain Shams University, Cairo, Egypt, the Nanotechnology
Center, Regional Center for Food and Feed, Agricultural Research Center
(ARC), Giza, Egypt, the Regional Center for Mycology and
Biotechnology, Al-Azhar University and the Molecular Services
Laboratory (H316), Research Center, October University for Modern
Sciences and Arts (MSA) during the period from 2012 to 2017.
Silver nanoparticles (AgNPs) are widely used in a growing number
of applications due to their unique properties Due to the intensive
commercial application of silver nanoparticles (AgNPs), risk assessment
of this nanoparticle is of great importance. There is a growing production
and application of silver nanoparticles (AgNPs) in various areas
including catalysis, consumer products, food technology, textiles/fabrics,
as well as medical products and devices. It was reported that about 25%
of the >1300 nanomaterial-containing consumer products contain
AgNPs. The rapid growth in the commercial use of AgNPs will
inevitably increase silver exposure in the environment and the general
population. However, with the accelerating introduction of AgNPs into
commercial products, there is likelihood of their release into the
environment, which gives rise to health and environmental concerns.
The aims of the present study were: 1.To investigates the impact
of AgNPs exposure in D. melanogaster with regard to changes in the
expression of heat shock (hsp) genes (hsp23, hsp26, hsp27, and hsp60).
2. To assess the correlations between hsps genes, antioxidant systems and
oxidative stress as reflected by changes in the expression of antioxidant
enzymes (SOD, CAT and GSH). 3. To assess the ability to tumors development by monitoring the changes in the expression of tumor
suppressor gene (p53). 4. To detect some genetic biomarkers associated
with biological stress in fruit flies.
The results could be summarized as the following:
(A) Physiochemical level
1- By using transmission electron microscopy (TEM): most of the
silver nanoparticles AgNPs were shown to be in a spherical shape
and with an average diameter of 15-70 nm.
2- By using dynamic light scattering (DLS): silver nanoparticles were
found to be mono dispersed with hydrodynamic diameter 74.35 nm.
(B) Toxicity results;
1- Larvae treated with six concentrations of silver nanoparticles (25,
100, 200, 400, 800 and 1600μg/ml), revealed a reduced larval and
pupal survival, affected larval pigmentation, adult loss of melanin,
defects in adults and the appearance of some malformations
compared with the control.
2- Silver nanoparticles AgNPs stimulated the mortality rate of -
Drosophila larvae. The average survival rate of the control (water)
was 98%.In concentrations of (25 μg/ml), (100 μg/ml), (200
μg/ml),(400 μg/ml),(800 μg/ml) and (1600 μg/ml) 4%,6% ,10%,
16% ,36%, and 50% of larvae were killed, respectively. This allowed
the determination of the value of “LC50” at a concentration of
1600μg / ml.
3- Different concentrations of silver nanoparticles during the larval
stage resulted in cuticle defects in adults. At AgNPs of 800 mg/L and
above, adult flies were appeared lighter in body color with little or
no melanin pigments left in their body. AgNPs selectively interferes
with the melanin pigmentation.
4- Increasing the concentration of Silver Nano-particles from 25 to1600
μg/ml inhibited cuticle sclerotization as indicated by the light color
of the larvae.5- Larvae exposure to 800 μg/ml AgNPs and above, effectively
caused melanization and pale abdomen of female in adult flies as
compared to wild type flies
6- Treatment with AgNPs revealed dramatic phenotypic modifications
in the subsequent generations of Drosophila resulted in production
of abnormal adults inducing leg and wing abnormalities as well as
deformations of wings, eyes, and thorax.
(C) Biochemical level
1- DPPH assay;
This assay was used to measure the activity of antioxidant molecules
and resulted in;
a) Decrease of antioxidant activity of larvae treated by silver
nanoparticles.
b) Increase of antioxidant activity of larvae treated by heat shock
compared to larvae treated with silver nanoparticles.
c) Decrease of antioxidant activity of larvae treated with silver
nanoparticles, followed by heat shock treatment compared to
treatment by heat shock only.
d) DPPH free radical scavenging assay revealed that AgNPs reduce the
antioxidant activity compared with control that may affect on the
immune system due to low antioxidant activity.
2- Reactive oxygen species (ROS) assay;
a) Larvae treated with silver nanoparticles showed an increase
amount of ROS.
b) Larvae treated with heat shock showed an increase of ROS
compared to the control larvae.
c) Larvae treated with silver nanoparticles followed by heat shock
treatment revealed a significant increase in ROS compared to the
control larvae.
(D) Molecular genetic analysis:
Experiments were carried out with the use of RT-PCR using 2X
SYBR® Green PCR master Mix assay to measure the relative
changes in mRNA expression levels using four hsps genes (hsp23,
hsp26, hsp27, and hsp60), three antioxidant genes (SOD,CAT and
GSH) and tumor-suppressor gene (p53) in order to:
1- Assess the correlations between hsps genes, antioxidant
genes and oxidative stress.
2- Assess antioxidant enzymes (SOD, CAT and GSH)
3- Assess tumor-suppressor gene (p53).
a) The results revealed significant changes in the expression of the
four hsp genes (hsp23, hsp26, hsp27, and hsp60) occurred in
larvae after exposure to 1600μg/ml AgNPs followed by heat
shock treatment.
b) Gene expression was determined as the relative fold change
normalized with RP49 mRNA expression. The expression of
hsp26, hsp23, and hsp27 genes transcription level in the heat
shock treatment were increased respectively (44.67-Fold),
(15.21-Fold), (5.56-Fold), while the expression of hsp60 gene
transcription level was decreased (0.29-Fold).The expression of
hsp23, hsp60, and hsp27 genes transcription level in the AgNPs
nano-stress treatment were decreased by (0.76-Fold), (0.53-
Fold), (0.37-Fold), respectively. The expression of hsp26 gene
transcription level was increased (4.65-Fold).While the
expression of hsp26, hsp27, hsp23 and hsp60 genes transcription
level in the AgNPs nano-stress+ heat shock treatment were
increased by (13.34-Fold), (1.32-Fold), (1.19-Fold),(1.06-Fold),
respectively.
c) Antioxidants and tumor-suppressor gene (p53).
The study revealed a significant increase in the gene
expression of antioxidant genes (CAT, SOD, GSH) and
tumor-suppressor gene (p53) as follows:
a) The levels of gene expression due to exposure to heat
shock were increased by (2.60-Fold), (2.14-Fold), (1.43-
Fold), (1.14-Fold), respectively.b) The expression of p53 and GSH genes transcription
levels in the AgNPs nano-stress treatment were
increased by (16.03-Fold), (1.34-Fold), respectively.
c) The levels of gene expression of SOD and CAT genes
decreased by (0.59-fold) and (0.42-fold), respectively.
d) The expression ofp53, GSH, CAT and SOD genes in
the AgNPs nano-stress heat shock treatment were
increased by (50.36-Fold), (1.84-Fold), (1.32-Fold)
and (1.29-Fold), respectively.
Finally, these results sound alarming; potential health and
environmental effects of nanoparticles need to be thoroughly assessed
before their widespread commercialization. Though there are few studies
on cytotoxicity of nanoparticles on mammalian and human cell lines,
there are hardly any reports on genotoxic and cytotoxic behavior of
nanoparticles in plant cells. They underline the importance of systematic
and reliable toxicology characterizations of nanomaterials and the
necessity of extensive efforts by the nano-science community in
designing and testing suitable nanoscale surface engineering/coating to
develop biocompatible nanomaterials with no hazardous effects for
human health and environment.