Induction Of Micronuclei Research Articles

Toxicity and Genotoxicity of Three Antimicrobials Commonly Used in Veterinary Medicine.

The toxicity of chlortetracycline (CTC), oxytetracycline (OTC) and enrofloxacin (ENF) was tested on two green algal species: the international standard Pseudokirchneriella subcapitata and the native Argentine species Ankistrodesmus fusiformis. All three antibiotics inhibited the algal growth. The most sensitive species was P. subcapitata, for which the EC50 for CTC, OTC and ENF were 1.19 ± 0.53, 0.92 ± 0.30 and 5.18 ± 3.80mg L-1, respectively. The EC50 for A. fusiformis, were 3.23 ± 0.53, 7.15 ± 2.69 and 10.6 ± 1.28mgL-1, respectively. The genotoxicity of these veterinary antibiotics was also assessed using chromosome aberration (CA) and micronuclei (MN) induction in Allium cepa roots. Three concentrations were tested (0.1, 1 and 10mgL-1). Only ENF at 1 and 10mgL-1 showed any significant MN induction. These data revealed that CTC, OTC and ENF could cause toxicity on green algae, whereas ENF could cause genotoxicity on A. cepa plants.

Toxicity of two effluents from agricultural activity: Comparing the genotoxicity of sugar cane and orange vinasse

Vinasse, produced by several countries as a by-product of agricultural activity, has different alternatives for its reuse, mainly fertirrigation. Several monocultures, such as sugar cane and orange crops, produce this effluent. Sugar cane vinasse is already widely used in fertirrigation and orange vinasse has potential for this intention. However, its use as a fertilizer has caused great concern. Thus, ecotoxicological evaluation is extremely important in order to assess the possible effects on the environment. Therefore, the aim of this study was to evaluate the potential toxicity of vinasse of two different crops: sugar cane and orange. For this purpose, bioassays with Allium cepa as a test organism were performed with two vinasse dilutions (2.5% and 5%) to detect chromosomal aberrations and micronucleus induction. The results showed that both types of vinasse are able to induce chromosomal aberrations in meristematic cells, mainly nuclear and anaphasic bridges, suggesting genotoxic potential. The induction of micronuclei in cells of the F1 region suggests that the two residues have mutagenic potential. Thus, caution is advised when applying these effluents in the environment.

Isoniazid Induced Toxicities and Idiosyncratic Responses in Male Albino Wistar Rats

Isoniazid (INH) is an anti-tuberculosis drug administered over a long period. Upon metabolism in the liver, INH generates nitrogen-centered radicals, reacting with cellular macromolecules, and induces toxic and transformational changes in cells and tissues. Here we examined the side effects of long-term (chronic) administration of isoniazid (2.5 and 5mg/kg) once daily for 30, 60 and 90 days consecutively: on hepatic transaminases, histological changes in hepatocytes and induction of micronuclei in the bone marrow and possible genotoxicity in E. coli PQ37. In addition, blood glucose was monitored during the various treatment period. Biochemical analysis of hepatic transaminases (γ-glutamyl-, alanine amino-, aspartate aminotransferases and alkaline phosphatase) in INH treated group was significantly (p<0.05) elevated as well as blood glucose level declined significantly (p<0.05) for both doses at 30, 60 and 90 treatment respectively. Total protein and albumin level decreased (p<0.05) at both treatment doses compared to control. Serum creatinine level significant (p<0.05) increased at days 30 and 60 relative to control.Antioxidants (GST, SOD and catalase) activity examined indicated a decrease in catalase levels which was significant (p<0.05) and an insignificant decrease (p>0.05) in GST in both treatment groups at day 60. There was also a significant increase (p<0.05) in the activity of superoxide dismutase activity. Micronucleus analysis further revealed an induction of micronucleated polychromatic erythrocytes (mPCEs), which was significant (p<0.05) for both treatment doses at days 30, 60 and 90 respectively. In addition, INH genotoxicity assessed by UMU chromotest indicated that the 5mg/kg dosage has an induction ratio above the genotoxicity threshold of 1.5 suggesting genotoxicity in E.coli PQ37.Taken together, INH treatment at both doses (2.5 and 5mg/kg body weight) was hepatotoxic and induced nephrotoxic damages, in addition to mutagenic effect which is more pronounced at 2.5mg/kg dose, thereby suggesting dose-dependent cellular and genetic toxicity.

Genotoxicity kinetics in murine normoblasts as an approach for the in vivo action of difluorodeoxycytidine.

This study analyzed the kinetics of in vivo micronucleus induction in normoblasts by determining the kinetics of difluorodeoxycytidine (dFdC)-induced micronucleated polychromatic erythrocytes (MN-PCEs) in the peripheral blood of mice. The kinetic indexes of MN-PCE induction of dFdC were correlated with the previously reported mechanisms DNA damage induction by this compound. In general, this study aimed to establish an in vivo approach for discerning the processes underlying micronucleus induction by antineoplastic agents or mutagens in general. The frequencies of PCEs and MN-PCEs in the peripheral blood of mice were determined prior to treatment and after treatment using dFdC at doses of 95, 190, or 380µmol/kg at 8h intervals throughout a 72h post-treatment. The area beneath the curve (ABC) for MN-PCE induction as a function of time, which is an index of the total effect, indicated that the dose response was directly proportional and that the effect of dFdC on micronucleus induction was reduced compared with that of aneuploidogens and monofunctional and bifunctional alkylating agents but increased compared with that of promutagens, which is consistent with our previous results. The ABC showed a single peak with a small broadness index, which indicates that dFdC has a single mechanism or concomitant mechanisms for inducing DNA breaks. The time of the relative maximal induction (T rmi) indicated that dFdC requires more time to achieve MN-PCE induction compared with aneugens and monofunctional and bifunctional alkylating agents, although it requires a similar time to achieve MN-PCE induction as azacytidine, which is consistent with evidence showing that both agents must be incorporated into DNA for their action to be realized. The timing of maximal cytotoxicity observed with the lowest dFdC dose was correlated with the timing of the main genotoxic effect. However, early and late cytotoxic effects were detected, and these effects were independent of the genotoxic response. A correlation analysis indicated that dFdC appears to induce MN-PCEs through only one mechanism or mechanisms that occur concomitantly, which could be explained by the previously reported concurrent inhibitory effects of dFdC on DNA polymerase alpha, polymerase epsilon, and/or topoisomerase. The timing of maximal cytotoxicity was correlated with the timing of maximal genotoxicity; however, an early cytotoxic effect that appeared to occur prior to the incorporation of dFdC into DNA was likely related to a previously reported inhibitory effect of dFdC on thymidylate synthase and/or ribonucleotide reductase.

Microelectrophoretic study of environmentally induced DNA damage in fish and its use for early toxicity screening of freshwater bodies.

This study investigates the potential of the comet and micronucleus assays of fish DNA as a means of screening the toxicity of aquatic environments. Catla catla and Cirrhinus mrigala collected from the River Chenab in Pakistan were used as a case study for the application of comet and micronucleus techniques. Comet and micronucleus assays were used to compare DNA damage in C. catla and C. mrigala collected from polluted areas of the River Chenab and farmed fish. Atomic absorption spectrophotometry showed an acute level of toxicity from Cd, Cu, Mn, Zn, Pb, Cr, Sn, and Hg in river water. Comet assay showed significant (p<0.05) DNA damage in C. catla representing 17.33±2.42, 11.53±2.14, and 14.17% DNA in the comet tail, averaged from three sites of the polluted area of the river. Tail moment was observed as 10.06±2.71, 3.11±0.74, and 14.70±1.89, while olive moment was 8.85±1.84, 3.83±0.76, and 7.11±0.73, respectively. Highly significant (p<0.01) damage was reported in C. mrigala as 37.29±2.51, 34.96±2.53, and 38.80±2.42% DNA in comet tail, tail moment was 23.48±3.90, 19.78±4.26, and 14.30±1.82, and olive moment was 16.22±2.04, 13.83±1.96, and10.99±0.90. Significant (p<0.05) differences were observed in genotoxicity between farmed and polluted area fish. Micronucleus assay showed a similar picture of significant difference in respect to single and double micronucleus induction: i.e., 23.20±4.19 and 2.80±1.07‰ in C. catla and 44.80±3.73 and 06.20±0.97‰, respectively, in C. mrigala. Nuclear abnormalities were found as 6.00±0.84 and 09.60±1.72/thousand cells, respectively, in both species. The results of this study suggest that these novel fish DNA damage assays can be used as an expedient toxicity screening for aquatic environments.

Evaluation of the potential of mobile phone specific electromagnetic fields (UMTS) to produce micronuclei in human glioblastoma cell lines

Some epidemiological studies indicate that mobile phones cause glioblastomas in humans. Since it is known that genomic instability plays a key role in the etiology of cancer, we investigated the effects of the universal mobile telecommunications system radiofrequency (UMTS-RF) signal, which is used in “smart” phones, on micronucleus (MN) formation and other anomalies such as nuclear buds (NBUDs) and nucleoplasmatic bridges (NPBs). MN are formed by structural and numerical aberrations, NBs reflect gene amplification and NPBs are formed from dicentric chromosomes. The experiments were conducted with human glioblastoma cell lines, which differ in regard to their p53 status, namely U87 (wild-type) and U251 (mutated). The cells were cultivated for 16h in presence and absence of fetal calf serum and exposed to different SAR doses (0.25, 0.50 and 1.00W/kg), which reflect the exposure of humans, in presence and absence of mitomycin C as former studies indicate that RF may cause synergistic effects in combination with this drug. We found no evidence for induction of MN and other anomalies. However, with the highest dose, induction of apoptosis was observed in U251 cells on the basis of the morphological features of the cells. Our findings indicate that the UMTS-RF signal does not cause chromosomal damage in glioblastoma cells; the mechanisms which lead to induction of programmed cell death will be investigated in further studies.

An in vitro study on the induction of micronuclei and other nuclear anomalies in peripheral blood lymphocyte culture by metal oxide nanoparticles

An in vitro study on the induction of micronuclei and other nuclear anomalies in peripheral blood lymphocyte culture by metal oxide nanoparticles

Radiosensitization by the ATR Inhibitor AZD6738 through Generation of Acentric Micronuclei.

AZD6738 is an orally active ATR inhibitor (ATRi) currently in phase I clinical trials. We found in vitro growth inhibitory activity of this ATRi in a panel of human cancer cell lines. We demonstrated radiosensitization by AZD6738 to single radiation fractions in multiple cancer cell lines independent of both p53 and BRCA2 status by the clonogenic assay. Radiosensitization by AZD6738 to clinically relevant doses of fractionated radiation was demonstrated in vitro using a 3D tumor spheroid model and, in vivo, AZD6738 radiosensitized by abrogating the radiation-induced G2 cell-cycle checkpoint and inhibiting homologous recombination. Mitosis with damaged DNA resulted in mitotic catastrophe as measured by micronucleus formation by live-cell fluorescent-ubiquitination cell-cycle imaging of cell-cycle progression and nuclear morphology. Induction of micronuclei was significantly more prominent for AZD6738 compared with inhibition of the downstream kinase CHK1 alone at isoeffective doses. Micronuclei were characterized as acentric chromosomal fragments, which displayed characteristics of increased DNA damage and cell-cycle dyssynchrony when compared with the primary nucleus. Mol Cancer Ther; 16(1); 25-34. ©2016 AACR.

Methanol Extract of Adansonia digitata Leaf Protects Against Sodium Arsenite-induced Toxicities in Male Wistar Rats.

Background:Human and animal population exposure to arsenic through the consumption of arsenic contaminated water is rampant in many parts of the world. Protective agents of medicinal plants origin could provide maximum protection against toxicities of various kinds.Objective:The protective role of orally administered methanol extract of the leaves of Adansonia digitata (MELAD) on sodium arsenite (SA) – induced clastogenicity and hepatotoxicity in male Wistar rats was evaluated.Materials and Methods:Thirty male Wistar rats divided into six Groups (1–6) of five animals each were used for the study. Group 1 (negative control) received distilled water and normal diet only, Groups 2–6 received the extract (at 250 or 500 mg/kg body weight) and/or SA at 2.5 mg/kg body weight.Results:There was statistically significant (P < 0.05) increase in the number of micronucleated polychromatic erythrocytes and lipid peroxidation in the SA group as compared with the negative control and treated groups. Administration of the extract reduced the effects of SA on the above parameters. Activities of serum alanine and aspartate aminotransferases did not show statistically significant effects; however, the histological analyses revealed periportal cellular infiltration by mononuclear cells, whereas the MELAD treated groups show mild cellular infiltration and mild portal congestion.Conclusions:MELAD protect against SA-induced toxicities in rats, and it may offer protection in circumstances of co-exposure and cases of arsenicosis.SUMMARY MELAD extract significantly reduce the lipid peroxidation induced by sodium arsenite in the liver of rats.MELAD did not show profound effects on the activities of serum alanine (ALT) and aspartate (AST) aminotranferases.MELAD offered significant protection against sodium arsenite-induced genotoxicity in the micronuclei induction assay.In the circumstances of co-exposure to arsenic contamination, MELAD may protect against sodium arsenite-induced toxicities. Abbreviations Used: MELAD: Methanol extract of the leaves of Adansonia digitata, SA: Sodium arsenite, nMPCEs: Number of micronucleated polychromatic erythocytes; ALT: Alanine aminotranferase; AST: Aspartate aminotranferase, TBARS: Thiobarbituric acid reactive substances, TBA: Thiobarbituric acid, MDA: malondialdehyde, Sodium arsenite (NaAsO2), IARC: International Agency for Research on Cancer

Aneugenicity and clastogenicity in freshwater fish Oreochromis niloticus exposed to incipient safe concentration of tannery effluent

Conventional effluent bioassays mostly rely on overt responses or endpoints such as apical and Darwinian fitness. Beyond the empirical observation, laboratory toxicity testing needs to rely on effective detection of prognostic biomarkers such as genotoxicity. Indeed, characterization of tannery effluent requires slotting in of genotoxic responses in whole effluent toxicity testing procedures. Hence, the prime objective of the present experimental investigation is to apply the technique of biological assay as a tool of toxicity testing to evaluate the induction of micronuclei (MN) in peripheral erythrocytes, and exfoliated cells of gill and kidney of O.niloticus exposed to Maximum tolerable concentrations (MTCs) of composite Modjo tannery effluent (CMTE) and to compare the sensitivity of each cells origin to the induction of MN. After 72h of exposure, cellular aberrations were detected using MN and nuclear abnormality (NA) tests. The induction of MN was significantly higher in exposed groups (P<0.05) when compared to the control group; moreover the tissue specific MN response was in the order, gill cells>peripheral erythrocyte>kidney. Total NA was found to increase significantly (P<0.05), when compared to the non-exposed group. NA was also further ramified as blebbed (BL), bi-nucleated (BN), lobbed (LB) and notched (NT) abnormalities. The result of each endpoint measured has demonstrated that at a concentration of total chromium (0.1, 0.73 and 1.27mg/L), a perceptible amount cellular aberration was measured, further implicating somber treat of genotoxicity to fishes, if exposed to water contaminated with tannery effluent. This further highlight that conventional effluent monitoring alone cannot reveal the effects expressed at cellular and genetic levels further demanding the incorporation of effluent bioassays in risk assessment and risk management/abatement programs.

The genotoxic potency of glycidol established from micronucleus frequency and hemoglobin adduct levels in mice

Glycidol is a genotoxic animal carcinogen that has raised concern due to its presence in food, as glycidyl fatty acid esters. Here we investigated the genotoxicity of glycidol in BalbC mice (0–120 mg/kg) by monitoring the induction of micronuclei in peripheral blood as a marker of chromosomal damage. The scoring of the micronuclei was assessed by flow cytometry. In the treated mice, the internal dose of glycidol, expressed as area under the concentration-time curve, AUC (mol × L−1 × h; Mh), was measured by dihydroxypropyl adducts to hemoglobin (Hb). The study showed that glycidol induced linear dose-dependent increases of Hb adducts (20 pmol/g Hb per mg/kg) and of micronuclei frequencies (12‰ per mMh).Compared to calculations based on administered dose, an improved dose-response relationship was observed when considering internal dose, achieved through the applied combination of sensitive techniques used for the scoring of micronuclei and AUC estimation of glycidol in the same mice. By comparing with earlier studies on micronuclei induction in mice exposed to ionizing radiation we estimated the radiation dose equivalent (rad-eq.) of glycidol to be ca 15 rad-eq./mMh.

Protective effect of mild endoplasmic reticulum stress on radiation-induced bystander effects in hepatocyte cells.

Radiation-induced bystander effect (RIBE) has important implications for secondary cancer risk assessment during cancer radiotherapy, but the defense and self-protective mechanisms of bystander normal cells are still largely unclear. The present study found that micronuclei (MN) formation could be induced in the non-irradiated HL-7702 hepatocyte cells after being treated with the conditioned medium from irradiated hepatoma HepG2 cells under either normoxia or hypoxia, where the ratio of the yield of bystander MN induction to the yield of radiation-induced MN formation under hypoxia was much higher than that of normoxia. Nonetheless, thapsigargin induced endoplasmic reticulum (ER) stress and dramatically suppressed this bystander response manifested as the decrease of MN and apoptosis inductions. Meanwhile, the interference of BiP gene, a major ER chaperone, amplified the detrimental RIBE. More precisely, thapsigargin provoked ER sensor of PERK to initiate an instantaneous and moderate ER stress thus defensed the hazard form RIBE, while BiP depletion lead to persistently destroyed homeostasis of ER and exacerbated cell injury. These findings provide new insights that the mild ER stress through BiP-PERK-p-eIF2α signaling pathway has a profound role in protecting cellular damage from RIBE and hence may decrease the potential secondary cancer risk after cancer radiotherapy.

Mutagenic, genotoxic and bioaccumulative potentials of tannery effluents in freshwater fishes of River Ganga

ABSTRACTThe tannery industries are the reason of major environmental concerns as they release toxic heavy metals, like chromium, in rivers posing risks of genotoxicity and mutagenicity in aquatic organism and indirectly in humans through food chain. In the present analysis, the freshwater inhabitant fishes of River Ganges, viz., Labeo calbasu, Puntius sophore, and Mystus vittatus, were examined for assessing the genotoxic, mutagenic, and bioaccumulative potentials of tannery effluents. For genotoxicity assessment, the blood and gill samples of fishes prevailed from polluted sites of River Ganges adjoining Kanpur city were utilized for comet assay and micronucleus test. The present investigation revealed the presence of significantly (p < 0.05) higher micronuclei induction and % tail DNA in erythrocytes and gill cells of the fishes collected from the polluted sites. The bioaccumulation studies revealed chromium concentration in muscle (0.89 µg/g) and gill tissues (0.24 µg/g) of L. calbasu; muscle (0.44 µg/g) and gills (1.23 µg/g) of P. sophore; and muscle (0.9617 µg/g) and gills (0.3628 µg/g) of M. vittatus, quite higher than the permissible limits of the World Health Organization. Consequently, the present study indicates strongly that River Ganges is contaminated with harmful tannery pollutants causing genotoxicity and mutagenicity in freshwater fishes.

Determination of heavy metal genotoxicity and their accumulation pattern in different fish organs of selected fish species collected from the Asa River, Ilorin, Kwara State, Nigeria

This study investigated bioaccumulation of heavy metals and cytogenotoxic effects that could result from exposure of fish to heavy metals in the Asa River, Ilorin, Nigeria. The three different fish species, Tilapia zilli, Oreochromis niloticus and Sarotherodon galilaeus were obtained from the Asa River and the fourth species, Clarias gariepinus was cultured in the laboratory to serve as control. The fish organs; bone, gills, kidney and liver from the four fish species were carefully dissected for the determination of some heavy metals. The results showed that the metal concentration levels were in the order Pb&gt; Cr &gt; Zn &gt; Cd. The highest lead concentration value was observed in the gills of Tilapia fish. The lowest level of Cd was observed in the bone tissue of Oreochromis niloticus. The values of heavy metals found in the cultured fish species were very low in concentration compared to the ones sourced from Asa River. Micronuclei and frequencies of nuclear abnormalities were assessed in the blood erythrocytes of both laboratory cultured fish and the ones obtained from Asa River. Significant differences (P&lt;0.05) were observed for mean frequencies of micronucleus induction in the blood erythrocytes of Oreochromis niloticus, Sarotherodon galilaeus. As for other nuclear abnormalities, significant differences (P&lt;0.05) were observed for mean frequencies in the blood erythrocytes, gills and liver of Tilapia zilli and Oreochromis niloticus. Genotoxicity results in this study showed that the Asa river water contains genotoxic metals say be responsible for the micronucleus and nuclear abnormalities observed in the aquatic environment.Keywords: Genotoxicity, bioaccumulation, organs, cultured, metals, fish organs

Evaluation of the cytotoxic and genotoxic effects of benchmark multi-walled carbon nanotubes in relation to their physicochemical properties

To contribute with scientific evidence to the grouping strategy for the safety assessment of multi-walled carbon nanotubes (MWCNTs), this work describes the investigation of the cytotoxic and genotoxic effects of four benchmark MWCNTs in relation to their physicochemical characteristics, using two types of human respiratory cells.The cytotoxic effects were analysed using the clonogenic assay and replication index determination. A 48h-exposure of cells revealed that NM-401 was the only cytotoxic MWCNT in both cell lines, but after 8-days exposure, the clonogenic assay in A549 cells showed cytotoxic effects for all the tested MWCNTs. Correlation analysis suggested an association between the MWCNTs size in cell culture medium and cytotoxicity.No induction of DNA damage was observed after any MWCNTs in any cell line by the comet assay, while the micronucleus assay revealed that both NM-401 and NM-402 were genotoxic in A549 cells. NM-401 and NM-402 are the two longest MWCNTs analyzed in this work, suggesting that length may be determinant for genotoxicity. No induction of micronuclei was observed in BBEAS-2Beas-2B cell line and the different effect in both cell lines is explained in view of the size-distribution of MWCNTs in the cell culture medium, rather than cell’s specificities.

Efeitos do Cromo Hexavalente Sobre o Crescimento de Raízes e Ciclo Celular no Meristema da Ponta da Raiz de Allium cepa

The International Agency Research on Cancer (IARC) classifies the hexavalent chromium [Cr (VI)] as carcinogenic to humans. The major source of environmental contamination by chromium in aquatic ecosystems is related to industrial application of this metal. In this study, we evaluated the cytotoxic and genotoxic effects of Cr (VI) in cell cycle meristem of Allium cepa root. The test was used to evaluate the effects of chromium on seed germination, root elongation, mitotic index and the induction of micronuclei and anomalies of the mitotic cycle. Allium cepa seeds were germinated in five different conditions: (1) negative control, distilled water; (2) test group T1, treated with potassium dichromate 6 mg.L -1 ; (3) test group T2, treated with potassium dichromate 12 mg.L -1 ; (4) test group T3, treated with potassium dichromate 24 mg.L -1 ; (5) positive control, treated with NMU (N-nitroso-N-methylurea) 0,125 mg.L -1 . The statistical analyses used were ANOVA and Pearson's correlation coefficient. Though without affecting the seed germination rate and elongation of roots, the results demonstrated cytotoxic effects on the mitotic index, as well as genotoxic effects of hexavalent chromium. Keywords : Chromium; Allium cepa ; cell cycle; cytotoxicity; genotoxicity.

Dose and Radioadaptive Response Analysis of Micronucleus Induction in Mouse Bone Marrow.

Enhanced cellular DNA repair efficiency and suppression of genomic instability have been proposed as mechanisms underlying radio-adaptive responses following low-dose radiation exposures. We previously showed that low-dose γ irradiation does not generate radio-adaptation by lowering radiation-induced cytogenetic damage in mouse spleen. Since radiation may exert tissue-specific effects, we extended these results here by examining the effects of γ radiation on cytogenetic damage and proliferative index in bone marrow erythrocytes of C57BL/6 and BALB/c mice. In C57BL/6 mice, the induction of micronuclei in polychromatic erythrocytes (MN-PCE) was observed at radiation doses of 100 mGy and greater, and suppression of erythroblast maturation occurred at doses of >500 mGy. A linear dose–response relationship for MN-PCE frequencies in C57BL/6 mice was established for radiation doses between 100 mGy and 1 Gy, with departure from linearity at doses of >1 Gy. BALB/c mice exhibited increased MN-PCE frequencies above baseline following a 20 mGy radiation exposure but did not exhibit radio-sensitivity relative to C57BL/6 mice following 2 Gy exposure. Radio-adaptation of bone marrow erythrocytes was not observed in either strain of mice exposed to low-dose priming γ irradiation (single doses of 20 mGy or 100 mGy or multiple 20 mGy doses) administered at various times prior to acute 2 Gy irradiation, confirming the lack of radio-adaptive response for induction of cytogenetic damage or suppression or erythrocyte proliferation/maturation in bone marrow of these mouse strains.

Comparative genotoxicity of silver nanoparticles in human liver HepG2 and lung epithelial A549 cells.

With the rapid expanding of human exposure to silver nanoparticles (AgNPs), genotoxicity screening of nanosilver is necessary to ensure consumer safety. Here, we assessed one key DNA damage responsive pathway activated by GADD45a gene after 24 h of AgNPs exposure in stable luciferase reporter cell systems based on two widely used in vitro cell models, human liver HepG2 and lung epithelial A549 cells. The comet assay and micronucleus test were also conducted to confirm the genetic damage induced by AgNPs. Our results showed that AgNPs produced a strong dose-dependent increase in transcriptional activation of GADD45a promoter indicated by luciferase activity accompanying by the significant decreases in cell viability. Surprisingly, in HepG2-luciferase cells, the relative luciferase activity was greater than 4.5× the control level after being treated with 200 μg ml-1 AgNPs. These results were generally in line with the positive and dose-dependent responses in cytotoxicity, DNA strand breaks indicated by Olive tail moment, tail DNA (%) and tail length, and chromosome damage indicated by induction of micronuclei, nucleoplasmic bridges, and nuclear buds. Additionally, compared with the A549-luciferase cells, the HepG2-luciferase cells seemed to be more susceptible to AgNPs as higher levels of genotoxicity were induced. We concluded that our GADD45a promoter-driven luciferase reporter gene cell system, together with the comet assay and micronucleus test, can be used as valuable tools for rapid screening of genotoxic potential of nanosilver. Copyright © 2016 John Wiley & Sons, Ltd.

Factors affecting the in vitro micronucleus assay for evaluation of nanomaterials.

A number of in vitro methodologies have been used to assess the genotoxicity of different nanomaterials, including titanium dioxide nanoparticles (TiO2 NPs) and silver nanoparticles (AgNPs). The in vitro micronucleus assay is one of the most commonly used test methods for genotoxicity evaluation of nanomaterials. However, due to the novel features of nanomaterials, such as high adsorption capacity and fluorescence properties, there are unexpected interactions with experimental components and detection systems. In this study, we evaluate the interference by two nanoparticles, AgNPs and TiO2 NPs, with the in vitro micronucleus assay system and possible confounding factors affecting cytotoxicity and genotoxicity assessment of the nanomaterials including cell lines with different p53 status, nanoparticle coatings and fluorescence, cytochalasin B, fetal bovine serum in cell treatment medium and different measurement methodologies for detecting micronuclei. Our results showed that micronucleus induction by AgNPs was similar when evaluated using flow cytometry or microscope, whereas the induction by TiO2 NPs was different using the two methods due to TiO2's fluorescence interference with the cytometry equipment. Cells with the mutated p53 gene were more sensitive to micronucleus induction by AgNPs than the p53 wild-type cells. The presence of serum during treatment increased the toxicity of AgNPs. The coatings of nanoparticles played an important role in the genotoxicity of AgNPs. These collective data highlight the importance of considering the unique properties of nanoparticles in assessing their genotoxicity using the in vitro micronucleus assay.

Attesting the efficiency of monitored natural attenuation in the detoxification of sewage sludge by means of genotoxic and mutagenic bioassays

A viable alternative to the use of sewage sludge (SS) would be using it as a reconditioner of agricultural soils, due to its high content of organic matter and nutrients. However, this solution may contaminate the soil, since SS may contain toxic substances. Monitored natural attenuation is a process that can be used in the decontamination of SS before its disposal into the environment. The effectiveness of the natural attenuation of a domestic SS was evaluated over 12 months by assays of Salmonella/microsome and micronucleus (MN) in human hepatoma cells (HepG2). Mutagenic activity was observed for the Salmonella strain TA 100, with S9, for the extracts from periods 0–6 months of natural attenuation. Genotoxic effects were observed in HepG2 cells, for 0 and 2 months, in almost all tested concentrations. Comparing obtained data by MN test to chemical analyses, it is possible to observe a coincidence between the induction of MN and the quantity of the m- and p-cresol, since these compounds were present in the initial SS and after 2 months of natural attenuation, decreasing their concentrations in samples from 6 to 12 months. The positive results obtained with Salmonella/microsome (from 6 months) suggest a combined action of other substances in SS. These results indicated that this SS, in the earlier periods tested, is potentially genotoxic and mutagenic and that its disposal can lead to severe environmental problems. Thus, the use of the studied SS as reconditioner requires pre-processing for over than 6 months of natural attenuation.