African Journal of
Biotechnology

  • Abbreviation: Afr. J. Biotechnol.
  • Language: English
  • ISSN: 1684-5315
  • DOI: 10.5897/AJB
  • Start Year: 2002
  • Published Articles: 12486

Full Length Research Paper

Cytogenetic and molecular assessment of some nanoparticles using Allium sativum assay

Mona S. Al-Ahmadi
  • Mona S. Al-Ahmadi
  • Department of Biology, College of Science, University of Imam Abdulrahman Bin Faisal, P. O. Box 1982, Dammam 31441, Kingdom of Saudi Arabia.
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  •  Received: 15 July 2019
  •  Accepted: 26 August 2019
  •  Published: 30 September 2019

 ABSTRACT

One of the primary objectives in agriculture is providing high-quality crops to consumers. Multiple techniques and methods are utilized to achieve this objective, including nanotechnology that depends on the use of very small materials, which will help in decreasing the amounts usually used with similar effects. Nanomaterials are used as fertilizers and also as component of nano-pesticides for plants. Despite their benefits, however, studies have noted their potential for cytotoxicity and genotoxicity. In this study, five nanoparticles (NPs) were tested to assess their effects on plants. The chromosomal aberration assay was used. The results showed that some NPs decreased the mitotic index (MI) significantly, which indicates the NPs’ potential cytotoxicity. In addition, different NPs’ treatments caused different types of chromosomal abnormalities e.g., chromosomes stickiness and disturbance of the metaphase and anaphase, lagging chromosomes, bridges, disturbed poles, micronuclei, s-metaphase, s-telophase, c- metaphase and bi-nucleus cells. All treatments had significant effects at p≤005. Treatments with NPs concentrations for 24 h affected the DNA content, AlO2 and Fe3O4 NPs’ increased the DNA content, while CeO2, TiO2 and Ag NPs’ decreased it. High concentrations of the tested NPs decreased the DNA content. The study results showed that CeO2 was the most harmful NP compared to the control and other NPs. Some types of chromosome abnormalities such as lagging chromosomes, bridge, and micronuclei indicate potential genotoxicity for these NPs. Despite of the positive effects, they also had negative side effects such as decreasing the MI and increasing the occurrence of different types of chromosomal abnormalities.

 

Key words: Cytotoxicity, genotoxicity, nanoparticles, mitotic chromosomes abnormalities.


 INTRODUCTION

Nanotechnology has been used in many fields. In agriculture, one of these technologies involves the use of different elements in nano sizes, which can give satisfactory results using a low amount of the element compared with its natural size. Nanomaterials are used in various applications such as plants protection, nutrition and of farm practices management due to their small size, high surface-to-volume ratio, and unique optical properties (Ghormade et al., 2011).
 
 
Nanoparticles (NPs) interact with plants, causing many morphological and physiological changes depending on their properties (Khodakovskaya et al., 2012). Chen and von Mikecz (2005) demonstrated that some NPs can enter cell nuclei and may directly affect the structure and function of the DNA genome. The efficacy of NPs depends on their concentrations, and these concentrations differ from plant to plant (Siddiqui et al., 2015). Also, NPs can have positive and negative impacts in higher plants (edible plants) and on their consumers in the food chain (Rico et al., 2011).
 
The minute size of NPs, smaller than cells and cellular organelles, allows them to penetrate those basic biological structures, disrupting their normal function (Buzea et al., 2007). Zheng et al., (2005) concluded that the up-take efficiency and effects of NPs on growth and metabolic function vary among plants. The concentrations of NPs affect processes like germination and plant growth. Babu et al., (2008) also suggested that the NPs’ size gives them free entry inside cells, where they can interfere in normal cell function. Landsiedel et al., (2009), Kovacic and Somanathan, 2010 and Siddiqui et al., (2015) suggested that the ability of NPs to penetrate cells easily allows them to affect the intercellular organelles and nucleic acids. NPs characteristics such as their small size, their shape and their large surface-area-to-mass ratio, and their propensity to cross cell barriers and their interaction with intercellular contribute to potential cellular and genetic toxicity caused by the induction of oxidative stress. Hunt et al., (2013) assessed the effects of nano silver on Caenorhabditis elegans by measuring the 8-OH guanine levels and found that the silver induced oxidative damage in DNA. A similar result was found by ÒªekiÒ« et al., (2017) in tomato plants. Cobalt oxide NPs were investigated by Faisal et al., (2016) to assess their effect on eggplant DNA. The results indicated that cobalt oxide NPs induced DNA strand breaks and apoptosis. Also, NPs cause chromosomal aberrations as several researchers have discussed in their study of these effects in higher plants (Kumari et al., 2009; Ghosh et al., 2010; Landa et al., 2012; Mukherjee et al., 2016; Debnath et al., 2018).
 
Higher plants are recognized as being excellent indicators of the cytogenetic and mutagenic effects of environmental chemicals. The study of these plants is also useful for detecting environmental mutagens indoors and outdoors. These plants are highly reliable bioassays for monitoring and testing for genotoxins because of their high sensitivity (Grant, 1999).
 
In this study, five NPs were tested to estimate their cytotoxicity and genotoxicity using a chromosomal aberration assay and to determine their effect on the DNA content of Allium sativum.


 MATERIALS AND METHODS

Tested materials
 
Table 1 shows NPs and different concentrations chosen depending on previous studies that found out that treatment with these concentrations had a positive effect on root length, yield and quality, biomass, and plant growth without serious harm on plants.
 
Sample preparations
 
A. sativum, common name (garlic) 2n = 16, gained from local markets were  used  as testing material. The loose outer scales and old roots were scraped and suspended in small beaker with distilled water.
 
Treatments
 
A. sativum were suspended in a small beaker (50 ml) with distilled water to encourage the root tips to grow until they reached 0.5 to 1 cm in length; they were then transferred to another beaker containing freshly prepared solutions of tested NPs, Aluminium oxide, Ferric oxide, Cerium oxide, Titanium oxide and Silver. Low concentrations (20 mg, 0.025 g, 0.012 g, 10 mg, 0.0005 mg) and high concentrations (40 mg, 5.9 g, 0.024 g, 20 mg, 0.001 mg) sequentially, and left for different periods of time (6, 16, and 24 h). One bulb of garlic was used for each treatment. The negative control was root tips treated with distilled water only, used as a qualified sample to compare for the effects of tested materials.
 
 
Slides preparation
 
The treated roots tips and negative control (untreated) were detached, fixed in freshly prepared 3:1 (v/v) ethanol alcohol: glacial acetic acid for 24 h. The root tips of A. sativum were hydrolyzed in 1N HCL at 60°C for 8 min. The root tips were then washed with distilled water several times and stained with 1% acetocarmin. Five temporary slides were prepared using the squash technique. Two root tips on each slide were examined for the effects of NPs on the mitotic index (MI). The same slides were analyzed for the types and frequencies of chromosomal abnormalities produced by the examined NPs.
 
DNA studies
 
Fisher bioreagents Sure-prep RNA/DNA/Protein purification kit was used to extract the genetic material from plant tissues following the instruction of HiPuraTM product. The plant DNA isolation was done using the CTAB method. The concentration of the isolated DNA was measured by Scan drop (Analytik Jena) device.
 
Scoring of slides and data analysis
 
Studying slides
 
The slides were viewed under light microscope (Phenix P H 50 DB047VU) using the 40X objective lens immersion. The demonstrative slides for each physical aberration were photo-graphed using Phenix micro Image analyzer Software 2008 EnV2, 2.
 
Mitotic index
 
On one slide for each treatment, a total of 2000 cells, were scored. The mitotic index (MI) was expressed as the number of dividing cells per total cells scored, as per the following equation:
 
Mitotic index (MI) = (Total number of dividing cell/Total number of cell examined) × 100
 
Cytotoxicity
 
The mitotic index of the treated cells was compared with that of the negative control sample.
 
Genotoxicity test
 
Chromosomal aberration per dose of each NP was examined; the percentage of cells with aberrations of each dose for each NP was scored and compared with that of the negative control as per the following equation:
 
Chromosomal aberration frequency (CF) = Total number of abnormal cell/Total number of normal cell
 
Statistical analysis
 
A two-way analysis of variance was used for determining the significance of difference at p ≤ 0.05 (SPSS 16.0 for Windows statistical package).
 

 


 RESULTS

Mitotic index (MI)
 
The NPs effects on the MI of A. sativum root tip cells
 
Table 2 and Figures 1 and 2 show the treatment results; it appears that the AlO2 NPs 20 and 40 mg concentrations decreased the MI after treatment for 24 h. This treatment was insignificant at p≥0.05, while treatment with a low concentration of Fe3O4 for 24 h was significant at p≤0.05. Also, CeO2 NPs decreased the MI after treatment with a low concentration for 6 h and high concentration for 24 h. This result was insignificant at p≥0.05. The TiO2 NPs treatment with a low concentration and a high concentration for 24 h decreased the MI. This result was significant at p≤0.05. Treatment with a high concentration of Ag NPs for 16 h decreased the MI. This result was significant at P≤0.05. Some treatments with a low concentration of NPs increased the MI. The AlO2 low concentration for 16 h, high and low concentrations of Fe3O4, also high and low concentrations of CeO2 for 16 h, and a low concentration of Ag for 6 h increased the MI compared to the control. These results were insignificant p≥0.05.
 
Chromosomal aberrations (CA)
 
Examining the cytological aberrations in plants is an excellent way to detect genetic hazards that environmental substances may pose (Grant, 1978).
 
Table 3a-b and figures 3, 4, and 5 show the types of abnormalities found in the mitotic chromosomes of A. sativum root tip cells after treatment with different concentrations of NPs.
 
All tested materials affected the chromosomes and increased chromosomal aberrations compared to the control, and the results were significant at p≤0.05. The most harmful concentrations were AlO2 NPs after treatment with a high concentration of 40 mg for 6 and 16 h (0.2) compared to the control (0.04), Fe3O4 NPs after treatment with a high concentration of 0.05 g for 6 h (0.2) and Ag NPs (0.2) after treatment with a high concentration 0.001 mg for 24 h compared to the control (0.02).
 
 
 
The types of chromosomal abnormalities scored after treatment with different NPs were chromosomal disturbance and stickiness during metaphase and anaphase, sticky telophase, chromosomes bridges during the anaphase and telophase, micronuclei, lagging chromosomes, star metaphase and star telophase, bi-nucleus  cells,   and   disturbed   poles  during  anaphase. Specific types of chromosome aberrations were scored after treatment with some NPs and they were C-metaphase, lagging chromosome in the telophase stage micronuclei, bi-nucleus cells, after treatment with AlO2 and Fe3O4 NPs, furthermore AlO2 NPs caused the formation of abnormal anaphase poles. Treatment with CeO2 NPs caused the formation of Star-metaphase, ring chromosome, C-metaphase and telophase bridge. The types of the chromosomal abnormalities scored after treatment with TiO2 NPs were S-metaphase, lagging chromosome   during    anaphase    stage,    S-telophase, bi-nucleus cells and micronuclei. Ag NPs produced lagging chromosome, S- anaphase, abnormal pole of anaphase stage, chromosomes bridges and bi-nucleus cells. Some types of chromosome abnormalities indicated the potential genotoxicity of tested NPs, e.g., micronuclei, lagging chromosomes, and the chromosome bridges during anaphase and telophase.
 
 
 
 
DNA content
 
Table 4, 5 and figures 6 and 7 show the effect of different concentrations of NPs (AlO2, Fe3O4, CeO2, TiO2, Ag) on DNA content after 24 h.
 
All the tested NPs affected the DNA content. Specifically, the content decreased after treatment with  low concentrations of Ag, TiO2 and CeO2 NPs (50.65, 55.32 and 97.63 ng/μl, respectively), and the results were significant at P≤0.05. AlO2 NPs and Fe3O4 NPs increased the DNA concentration (391.34 and 234.07 ng/μl, respectively) and these results were significant at   P≤0.05 compared to the control (144.73 ng/µl). Treatments with high concentrations affected the DNA content. The NPs Fe3O4, Ag and CeO2 (130.37l, 124.65, 119.33 ng/µl, respectively) decreased the DNA concentration, and these results were significant at p≤0.05; the results showed that CeO2 NPs were the most harmful and that TiO2 NPs were the least harmful  followed by  AlO2 compared to the control (144.73 ng/µl).
 
 
 


 DISCUSSION

Mitotic index (MI)
 
The NPs treatments reduced the MI. The decrease of MI might have resulted from the effect of  the  NPs  during  S-phase  which  inhibited  the DNA synthesis. The decrease might also be due to the activation of enzymes by decreasing or inhibiting the enzymes, particularly the enzymes that involved in DNA replication or cell division (Sudhakar et al., 2001).
 
AlO2 NPs caused decreased MI. This effect may be due to the blockage at GI stage, which disturbs the DNA synthesis (Mohandas and Grant, 1972). A similar result was found by Rajeshwari et al., (2015).
 
The effect of Fe3O4 NPs on cells was as reported by  Alarifi  et  al.,  (2014), that is, the cell death mediated by the reactive oxygen species (ROS) triggered mitochondrial pathway as evidenced by the cleavage of caspase-3 activity and caused an imbalance between the production and degradation of ROS and induced oxidative stress. NPs may change the production of ROS and affect antioxidation defense and so induce oxidative stress (Srinivas et al., 2011). More explanations of iron oxide reaction were reported by Zhongwen et al., (2012), that the cytotoxicity ability of iron oxide, iron oxide trapped in acidic lysosomes of   the    cell,    and     they    catalyze decomposition of H2O2 to produce hydroxyl radicals through peroxidase-similar activity.
 
The cytotoxicity of CeO2 may be due to the oxidative stress (Jezek and HIavata, 2005). Park et al., (2008) found that CeO2 caused cytotoxicity because of the introduction of ROS, and that the free radical species produced by CeO2 NPs significantly reduce the levels of cellular antioxidants. Also, Sendra et al., (2016) suggested that the toxicity of CeO2 NPs may be due to their photocatalytic properties. Similar results were demonstrated by Liman et al., (2019).
 
The TiO2 NPs decreased the MI compared to the control. Pakrashi et al. (2014) found that TiO2 NPs increased ROS and that this was the main contribution to the toxic effects. Castiglione et al., (2011) produced similar results in a study of the effect of TiO2 NPs on Vicia faba and Zea mays, while Klien and Godnic (2012) in a study of the effect of TiO2 NPs on rodents.
 
Ag NPs decreased the MI compared to the control. Patlolla et al., (2012) explained that the decrease in MI after treatment with different concentrations  of Ag NPs might be due to a lower progression of cells from S-phase to M-phase of the cell cycle. Babu et al., (2008) suggested that Ag NPs might affect the DNA synthesis during the S-phase cell cycle, leading to mitodepressive effects and cytotoxicity. These NPs might also cause slower development of cells from the S-phase (DNA synthesis) to the M-phase (mitosis) of the cell cycle as a consequence of silver NPs exposure (Kumari et al., 2009). Similar results were found by Pulate et al. (2011).
 
Some treatments had no effect on the MI while others increased it. This variance might be due to the intrinsic plant detoxification mechanism of NPs when the plants are exposed to nanotoxicity. Free metal radicals, formed during oxidative stress, function as signaling molecules that later activate the ROS detoxification and antioxidant  defense mechanisms in plants to deal with NPs toxicity (Zia-ur-Rehman et al., 2018).
 
Chromosomal aberrations (CA)
 
Treatments with different concentrations of NPs cause several types of chromosomal aberrations. Rajeshwari et al., (2015) found that AlO2 NPs decreased the MI and increased the chromosomal aberration in root cells of Allium cepa due to the ROS generated by the interaction of AlO2 NPs and root-tip cells.
 
The effects of Fe3O4 NPs were explained by Rajiv et al., (2015). They found that the metal-oxide NPs caused DNA damage and chromosomal aberrations due to the generation of ROS, which leads to cell death.
 
CeO2 NPs also produce chromosomes abnormalities. In this respect, Benameur et al., (2015) demonstrated that chromosomal aberrations are consistent with cellular ROS production. Similar result was found by Liman et al., (2019).
 
Treatment of A. sativum with different concentrations of TiO2 NPs for different time periods causes different types of chromosomal abnormalities; Ghosh et al., (2010) concluded that treatment with TiO2 NPs caused chromosomal aberration due to the generation of superoxide radicals that sequentially resulted in lipid peroxidation in the cells. Trouiller et al., (2009) found that TiO2 NPs are capable of causing oxidative bursts, resulting in DNA damage and the occurrence of micronuclei. Tavares et al. (2014) have the same effect of TiO2 NPs in human lymphocytes.
 
Ag NPs also cause chromosomal abnormalities. Kumari et al., (2009) suggested that Ag NPs could penetrate plant system and may impair stages of cell division, causing chromosomal aberrations. Similar results were found by Pulate et al. (2011) and Patlolla et al. (2012).
 
The presence of disturbance, S-metaphase, S-anaphase, S-telophase, lagging chromosomes, abnormal anaphase poles, and sticky chromosomes of metaphase and telophase revealed that NPs affected spindle fibers. Several studies concluded that NPs cause chromosomal aberration by affecting the spindle fibers. These aberrations alter the direction of chromosomes during different stages of mitotic division. This may be due to the interaction of NPs with mitotic spindle  apparatus,  centrioles  or  their  associated  proteins  leading  to  the  loss or gain  of chromosomes in daughter cells (Kuriyama and Sakai, 1974; Babu et al., 2008; Magdolenova et al., 2014).
 
The formation of chromosome stickiness involves the matrix of chromatin material which makes the chromosome stick or clump (Patil and Bhat, 1992). Klasterska et al., (1976) suggested that the stickiness of chromosomes arises due to the effect of NPs on nucleic acids, which causes polymerization and chromosomes stickiness. The formation of chromosomes bridges during anaphase and  telophase  may  be  due  to  chromosomal stickiness (EL-Khodar et al., 1990). Micronuclei being acentric fragments appear because of DNA breaks, especially during cell division, or because of laggards being excluded from the nucleus (Ma, 1982). These micronuclei could be owing to the inhibition of DNA synthesis at the S-phase (Kumari et al., 2009).
 
Grant (1978) reported that binucleate cells rise as a consequence of the inhibition of cell-plate formation. Huang et al., (2009) reported that due to the disruption of the mitotic checkpoint, PLKI protein function controls the mitosis process, including cytokinesis, when exposed to TiO2 NPs.
 
DNA content
 
Different treatments of NPs affect the DNA content. Kwon et al., (2014) suggest that small NPs cross the cellular membranes more easily and this can increase the potential for DNA damage. Within cells, many NPs end up in the lysosomes but some also appear in the cytoplasm and other cellular organelles, e.g., the Golgi body, the mitochondria, and the nucleus (Yuliang et al., 2010). The molecular mechanisms of NPs mostly depend on their chemical properties. Auffan et al., (2009) concluded that chemically stable metallic NPs have no significant cellular toxicity, while NPs that can be oxidized, reduced, or dissolved are cytotoxic and genotoxic for cellular organisms. Mehrian and Lima (2016) and Brunner et al., (2006) suggested three mechanisms involved in NPs toxicity. The first is the toxic substance from soluble NPs released into exposed media. These substances could contribute to DNA damage by their involvement in ROS generation (Fenton-type reaction) (Kruszewski et al., 2011). The second mechanism is the ROS generated through surface interactions with the media. The third mechanism is the direct physical interaction of NPs with biological targets such as cell membranes or DNA (Brunner et al., 2006). NPs can also interact with the mitochondria and other cell components and disrupt their functions. The ROS that result from the transfer of electrons’ energy to oxygen are highly reactive and potentially harmful to living organisms (Wu et al., 2014). Van Breusegern and Dat (2006) reported that ROS as a result of NP interaction will interact with almost all cellular components, producing protein change, lipid peroxidation, and DNA damage.
 
In this study, the treatment of A. sativum with AlO2 showed that a low concentration increases DNA content and a high concentration decreases it. Sjorgen and Larsen (2017) suggested that Al2 inhibits the cells’ entrance into the S-phase during the cell cycle, which will affect DNA content by decreasing the content frequency. On the other hand, the S-phase cells entered the G2/M phase, leading to an increase of DNA content frequency. Similar results were found by Silva et al., (2000) and Jaskowiak et al., (2018). Wu et al. (2014) demonstrate that the  reductive dissolution of iron oxide NPs induced a more homogeneous Fenton reaction, one that is more efficient in producing ROS. The availability of ROS inside the cell will affect cell components, and one of these components is DNA.
 
 
The  CeO2   NPs   was   the   most   harmful    for   DNA compared to other NPs and the control. This effect may be due to the instability of DNA caused by increasing oxidative stress, which leads to DNA damage that occurs due to the high presence of ROS (Mattiello et al., 2015). A   high   concentration   of   CeO2    NPs   effected   DNA content compared to the control but less than a low concentration, which has a greater impact on DNA content. This difference may be due to the superoxide dismutase (SOD) mimetic activity related to a high concentration of CeO2 NPs, which causes the dismutation of superoxide anions into H2O2 (Mattiello et al., 2015). Vranová et al., (2002) suggested that the oxidative burst induced by the more harmful dose of CeO2 NPs may be associated with the stimulation of cellular respiration that increases the signal requirement for energy. Mattiello et al., (2015) found that CeO2 NPs affect the DNA by inducing visible modifications in the chromatin aggregation. A condensed chromatin is a part of the programmed cell death. A similar result was found by Liman et al., (2019) for the effect of CeO2 NPs on the DNA content of A. cepa, by Benameur et al., (2015) for the effect on human dermal fibroblasts, and by Kumari et al., (2014) for the effect on Wistar rats. López-Moreno et al., (2010) found that CeO2 NPs affect the integrity of DNA and genetic stability of soybean plants.
 
In this study, the TiO2 NPs decreased the DNA content. As Rico et al., (2011) reported, this is due to the generation of superoxide radicals that cause lipid peroxidation in cells. Turkez and Geyikoglu (2007) reported that TiO2 NPs could induce genotoxicity by inducing sister chromatid exchange and micronuclei in human white blood cells. Also, Ghosh et al., (2010) reported that the effect of TiO2 NPs on DNA is due to the increased malondialdehyde (MDA) concentration that leads to lipid peroxidation, which leads to DNA damage. Pesnya (2013) concluded that TiO2 NPs have a high potential to interact with DNA and cause primary DNA damage. The bio-uptake effect of TiO2 NPs was explained by Pakrashi et al., (2014). They found a conjunction between the NPs uptake and the increase of ROS. An imbalance in intracellular ROS content caused by NPs exposure can induce DNA damages through oxidative stress owing to the oxidation of purine molecules (Afaq et al., 1998). Ghosh et al., (2012) found that treatment with TiO2 NPs caused genotoxicity because of the generation of superoxide radicles. Schins and Knaapen (2007) suggested that the genotoxic effect of TiO2 NPs might be due to oxidative stress and that the mechanism for this, as described by Donaldson et al., (1996) and Gilmour et al., (1997), is that TiO2 NPs have hydroxyl radical activity. Similar results for TiO2 NPs effect were found by Pakrashi et al., (2014).
 
Treatment of A. sativum with low and high concentrations of Ag NPs decreased the DNA content compared to the control. The Ag NPs induced toxicity due to their effect on ROS formation (Qian et al., 2013). Ma (1982) and Grant (1982) suggested that Ag  NPs   and  their   role  in  oxidative  stress  induced cellular death. Similar results were found by Sudhakar et al., (2001) and Babu et al., (2008). In higher plants, Saha and Gupta (2017) found that Ag NPs enter the plant cells and interfere with DNA repair, which leads to a blockage of DNA synthesis. Huijing et al., (2015) found that Ag NPs inhibit the new DNA synthesis in bacteria cells, which causes cell apoptosis.
 
This study showed that low concentrations of tested NPs had different effects on DNA. The Ag, TiO2 and CeO2 NPs decreased the DNA content, while AlO2 and Fe3O4 NPs increased it. This difference may result from the ROS generation (Mcshan et al., 2014). Sharma et al. (2012) reported that ROS’ destructive role depends on the equilibrium between ROS production and scavenging, that is, if a cell has developed a strong mechanism to control the ROS level by producing the enzymatic and non-enzymatic molecules needed to cope up with NPs-caused stress, it will decrease the effect of NPs on cell components including DNA.
 
The genotoxicity of NPs may result from their direct interaction with DNA or from indirect effects such as interacting with cells or tissues and releasing factors that cause harmful effects such as inflammation and oxidative stress (Singh et al., 2009; Magdolenova et al., 2014). Golbamaki et al., (2015) proposed that the genotoxic effects of NPs may be classified as primary genotoxicities or secondary genotoxicities. The second class may be due to the ROS generated during particle-induced inflammation, whereas the first class can be genotoxic without inflammation.
 
This study has revealed that different concentrations of the tested NPs affects the MI and that some treatments were significant at p ≤ 0.05 particularly, Fe3O4 NPs after treatment with a low concentration for 24 h, TiO2 NPs after treatment with low and high concentrations for 24 h, and Ag NPs after treatment with a high concentration for 16 h. This effect may be due to the free radicals generated by the interaction between NPs and cell components that raises the potential for cytotoxicity and decreases the MI. The tested NPs caused different types of chromosomal aberrations. Some of the scored types, e.g., micronuclei, lagging chromosomes, and chromosome bridges, indicated a genotoxic effect of NPs because these types of chromosome aberrations only occur if there is a direct effect on DNA. These NP effects may also be due to the time of interaction between the NPs and the cell cycle periods. It seems that NPs have greater effects during the S-phase of the cell cycle and wither this interaction starts during the beginning, middle, or end of the S-phase.


 CONFLICT OF INTERESTS

The authors have not declared any conflict of interests.



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