Tail Intensity Research Articles

Radiobiological effects of the alpha emitter Ra-223 on tumor cells

Targeted alpha therapy is an emerging innovative approach for the treatment of advanced cancers, in which targeting agents deliver radionuclides directly to tumors and metastases. The biological effects of α-radiation are still not fully understood - partly due to the lack of sufficiently accurate research methods. The range of α-particles is <100 μm, and therefore, standard in vitro assays may underestimate α-radiation-specific radiation effects. In this report we focus on α-radiation-induced DNA lesions, DNA repair as well as cellular responses to DNA damage. Herein, we used Ra-223 to deliver α-particles to various tumor cells in a Transwell system. We evaluated the time and dose-dependent biological effects of α-radiation on several tumor cell lines by biological endpoints such as clonogenic survival, cell cycle distribution, comet assay, foci analysis for DNA damage, and calculated the absorbed dose by Monte-Carlo simulations. The radiobiological effects of Ra-223 in various tumor cell lines were evaluated using a novel in vitro assay designed to assess α-radiation-mediated effects. The α-radiation induced increasing levels of DNA double-strand breaks (DSBs) as detected by the formation of 53BP1 foci in a time- and dose-dependent manner in tumor cells. Short-term exposure (1–8 h) of different tumor cells to α-radiation was sufficient to double the number of cells in G2/M phase, reduced cell survival to 11–20% and also increased DNA fragmentation measured by tail intensity (from 1.4 to 3.9) dose-dependently. The α-particle component of Ra-223 radiation caused most of the Ra-223 radiation-induced biological effects such as DNA DSBs, cell cycle arrest and micronuclei formation, leading ultimately to cell death. The variable effects of α-radiation onto the different tumor cells demonstrated that tumor cells show diverse sensitivity towards damage caused by α-radiation. If these differences are caused by genetic alterations and if the sensitivity could be modulated by the use of DNA damage repair inhibitors remains a wide field for further investigations.

Influence of Polymorphisms of DNA Repair and GST Genes on Genotoxic Damage and Mutagen Sensitivity in Workers Occupationally Exposed to Very Low Doses of Ionizing Radiation

The study investigated the influence of genetic polymorphisms of the enzymes for DNA repair and detoxification of reactive intermediates on spontaneous and bleomycin-induced (BLM) genotoxic damage in 43 workers exposed to very low doses of ionizing radiation (IR) (mean cumulative dose 5.31 mSv) and 43 subjects with no occupational exposure to IR (controls). In all the subjects examined, the frequency of chromosome aberrations (CAs) and micronuclei (MN), both spontaneous and BLM-induced, the Comet assay parameters (tail intensity), the genotypic variants of the DNA repair enzymes XRCC1 (Arg194Trp, Arg280His, Arg399Gln), XRCC3 (Thr241Met), XPD (Lys751Gln), and of the detoxification enzymes GSTM1 and GSTT1 (null genotype) and BLMH (A1450G) were determined. Among the biomarkers considered, only the frequency of total CAs (p &lt; 0.05), and in particular of chromosome breaks (p &lt; 0.01), was found to be significantly higher in the exposed workers than the controls. The frequency of spontaneous MN was higher in subjects with at least one allelic variant in XRCC1 than in carriers of the wild-type, but again only in exposed workers (p = 0.046). Linear regression analysis showed a positive dependency of the frequency of spontaneous chromosome breaks on occupational exposure, and a dependency of the frequency of BLM-induced MN negative on occupational exposure and positive on alcohol consumption and the null GSTM1 genotype. In conclusion, the frequency of chromosome breaks seems to be a useful cytogenetic biomarker for exposure to very low doses of IR, while only the combined effect of different gene variants or genetic, occupational, and lifestyle habits factors seems to be able to modulate the genotoxic effect of very low doses of IR.

DNA damage detection by Comet Assay on Daphnia magna: Application in freshwater biomonitoring

Monitoring of water genotoxicity still remains underexploited in risk assessment. The present study aimed at standardizing and evaluating the sensitivity and applicability of the Comet Assay adapted for Daphnia magna in genotoxicological investigations in freshwater environments. Two sampling campaigns (2014–2015) were performed in the watercourses of a pilot basin located in the Parma district (Italy). Fourteen sampling stations with different Ecological Status and/or EBI values were selected, all with a good Chemical Status according to the EU–Water Framework Directive 2000/60. The Alkaline Comet Assay was performed on 48 h-aged daphnids exposed (24 h) to 23 water samples. In parallel, the acute toxicity test was carried out. Daphnids exposed to samples, collected upstream the main watercourses in non-impacted areas, showed low DNA migration (Tail Intensity percentage – TI% - in the range 2.97–13.21), similar to laboratory controls. An increase in genotoxicity (TI% in the range 20–40) proceeding from the mountain towards the plain area was observed, in agreement with the land uses and the ES/EBI values of the stations. The highest genotoxic damage was observed after exposure to samples from watercourses of the minor hydrographic network in the plain area and waterbodies receiving wastewater treatment plant outflows. A modified version of the Comet Assay able to identify the presence of genotoxins inducing DNA oxidative damage, after standardization, was applied to daphnids treated with waters from 4 selected monitoring stations. The presence of oxidative contaminants was detected downstream a wastewater treatment plant outflow.The Comet Assay on D. magna has proven to be sensitive and able to discriminate among differently impacted areas and might be applied routinely. The FPG-Comet proved to be able to highlight the presence of contaminants causing oxidative stress.In our knowledge, this is the first time that Comet Assay on Daphnia magna is successfully applied for freshwater monitoring.

DNA damage in kidney and parenchymal and non-parenchymal liver cells of adult Wistar rats after subchronic oral treatment with tembotrione.

DNA damage in the liver and kidney cells of adult male Wistar rats was studied using the comet assay after a 28-day oral administration of tembotrione at doses of 0.0007, 0.0013 and 0.7 mg/kg b.w./day [AOEL (acceptable operator exposure level), REL (residual exposure level) and 1000× AOEL]. As a descriptor of DNA damage, tail intensity was used. Antioxidant status was assessed by activity of glutathione peroxidase (GPx). Significant DNA damage was recorded in the kidney cells at all three doses as compared to negative control. In parenchymal liver cells, significant DNA damage was observed in AOEL and 1000× AOEL doses, while in non-parenchymal liver cells, only AOEL-treated group was significantly different compared to negative control. In both types of liver cells, REL and 1000× AOEL doses were significantly different from the AOEL dose. No significant changes in GPx activity compared to control were observed at any exposure level. The results of the present study suggest that repeated in vivo exposure to tembotrione led to low-level DNA instability in kidney and liver cells. Exposure to the highest tembotrione dose showed a relatively weak response with the alkaline comet assay. Further research should focus on the effects of this herbicide in other models along with different exposure scenarios.

Oxidative Stress and Genotoxicity in Pterygium: A Systemic Investigation.

To perform a systemic investigation on oxidative stress and DNA damage in patients with primary pterygium. This prospective cross-sectional study included 32 patients with primary pterygium (60.1±2.0 years of age) and 33 age- and sex-matched (58.8±2.2 years of age) control subjects (P>0.05). A commercial kit was used for measuring serum total oxidant status (TOS) and total antioxidant status (TAS). The comet assay was performed after lymphocyte isolation from venous blood to quantitate DNA damage. Tail length (TL), tail intensity (TI), and tail moment (TM) were used for statistical analysis as parameters of DNA damage. In the pterygium group, TOS and TAS were significantly higher when compared with those of the control group (P=0.019 and P=0.005, respectively). In terms of DNA damage, patients with pterygium had higher TL, TI, and TM than in the control subjects (P<0.0001 for all). Although current literature focuses on local factors in pterygium pathogenesis, patients with pterygium seem to have increased systemic oxidative status (and compensatory antioxidant response) and genotoxicity, which might create a predisposition for pterygium development.

Photoautotrophically Grown Chlorella vulgaris Shows Genotoxic Potential but No Apoptotic Effect in Epithelial Cells.

This study investigated the effect of Chlorella vulgaris (C. vulgaris) on genotoxicity, cytotoxicity, and apoptosis in Caco-2 and HT-29 cells. C. vulgaris significantly induced DNA damage in both cell lines at a concentration of 200 μg dry matter/mL (comet tail intensity CTI: 24.6 ± 4.7% for Caco-2, 16.6 ± 0.9% for HT-29). The application of processing (sonication, ball-milling) did not affect the genotoxicity negatively and lowered the lipid peroxidation in C. vulgaris preparations. C. vulgaris-induced intracellular formation of reactive oxygen species in human cell lines and might be responsible for the genotoxic effect. A solid fraction mainly triggered the observed DNA damage (CTI: 41.5 ± 1.9%), whereas a hydrophilic (CTI: 7.9 ± 1.7%) and lipophilic (CTI: 10.2 ± 2.1%) fraction revealed a significantly lower tail intensity. C. vulgaris significantly induced DNA damage in both cell lines possibly through intracellular formation of reactive oxygen species; however, it was repaired after a 2 h recovery time or was even avoided at lower concentrations. In addition, none of the preparations indicated an adverse effect on cell proliferation or revealed apoptotic activity.

The comet assay in Ceraeochrysa claveri (Neuroptera: Chrysopidae): A suitable approach for detecting somatic and germ cell genotoxicity induced by agrochemicals

Some agrochemicals are genotoxic to several organisms. Nevertheless, few protocols are currently available for measuring the toxicogenetic effects of these compounds in target and non-target field-collected species of insects important to agriculture. Herein, we used the species Ceraeochrysa claveri (Neuroptera: Chrysopidae), a non-target predator insect, to investigate the ability of an azadirachtin-based biopesticide (Azamax™) to induce DNA damage. The alkaline version of the comet assay was standardized to evaluate genetic instability caused by the toxicant in somatic (gut) and germ (nurse cells and oocytes) cells of C. claveri. For this, C. claveri larvae were distributed into three groups (10/each) and treated with Azamax™ at 0, 0.3% or 0.5% throughout the larval stage. DNA damage (tail intensity) was measured in adult insects, four days after emerged. The data showed that both doses of Azamax™ (0.3% and 0.5%) were able to significantly (p < 0.05) increase DNA damage in somatic and germ cells of C. claveri. In conclusion, C. claveri (intestinal and ovarian cells) was a sensitive bioindicator for identifying Azamax™ genotoxic potential, whereas the comet assay was a useful tool for detecting the genotoxic hazard of the pesticide in the field-collected insect species. Given that estimation of adverse effects of pollutants on ecosystems is an essential component of environmental risk assessment, the approach used can be recommended to estimate the ecotoxicity of agricultural chemicals.

Genotoxic evaluation of polystyrene microplastic

Pollution of the aquatic environment by microplastic could be having a massive impact on marine life. As far as the dimensions of the microplastics decrease, the negative effects are also increasing. In this study, the effects of 1 µm diameter polystyrene microplastics (PSMs) on Daphnia magna and Neocaridina davidi were investigated. The acute toxicity test was conducted on Daphnia magna. According to the test LC50 value was calculated as 808.97 μg/mL. According to genotoxic evaluation on Neocaridina davidi with single cell gel electrophoresis (Comet), tail length, tail intensity and tail moment were increased by PSMs compared to the control.

Short‐term “in vivo” study on cellular DNA damage induced by acrylic Andresen activator in oral mucosa cells

To analyse through comet assay and micronucleus test the viability and DNA damage occurred in buccal mucosa epithelial cells after a short-term exposure to Andresen activator resin monomers. Test group consisting of 26 subjects was treated with Andresen activator; 16 subjects who had never undergone orthodontic treatment were enrolled in the control group. Buccal mucosa samples were collected before treatment and after 7, 15, 30, 60 and 90days. The analyses performed on the cells included the following: cellular viability, comet assay and micronucleus test. Mean±SD were calculated for cellular viability, tail moment, tail intensity, tail length, micronuclei, binuclear and bud cells. Significance (P<0.05) was evaluated with Dunnett's test. Cellular viability did not change during observational time, and its trend was similar to the controls. Tail moment and tail intensity significantly increased after 30 and 60days, respectively, whereas tail length remained unchanged over time in the test group; the same parameters did not change in the control group. In the test group, micronuclei, binuclear and bud cells significantly increased after 30, 60 and 90days, respectively. The resin monomers of the Andresen activator cause genotoxic effects detectable through comet assay and micronucleus test, but they do not produce clear cytotoxic effects after a 90days exposure.

DNA damage in rat kidneys and liver upon subchronic exposure to single and combined ochratoxin A and citrinin

The study aimed to check whether ochratoxin A (OTA) and citrinin (CIT) increase DNA damage in the kidney and liver of male Wistar rats (alkaline comet assay), clarify the oxidative nature of DNA damage (hOGG1-modified comet assay), and verify whether resveratrol (RSV) could ameliorate OTA+CIT-induced genotoxicity. Rats were treated orally with OTA (0.125 and 0.250 mg/kg bodyweight (bw)) and CIT (2 mg/kg bw), OTA+CIT combinations and OTA+CIT+RSV (0.250+2+20 mg/kg bw) for 21 days. Both alkaline and hOGG1-modified comet assay showed that DNA damage was more severe in rat kidneys than in liver following mycotoxin treatment. Alkaline comet assay revealed a higher intensity of DNA damage, particularly as measured by tail intensity in the kidneys. Both tail length and tail intensity were OTA dose-dependent, but in combined OTA+CIT treatment these values were similar to CIT alone and lower than in animals treated with single OTA, possibly due to induction of apoptosis. hOGG1-modified comet showed that OTA+CIT evoked greater oxidative DNA damage than single mycotoxins. RSV did not reduce DNA damage measured by alkaline comet assay, but hOGG1-modified comet showed that RSV ameliorated OTA+CIT genotoxicity in the kidneys. Apart from oxidative stress, other mechanisms of DNA damage are involved in OTA and CIT genotoxicity. In rat kidneys RSV can reduce but not overcome oxidative DNA damage induced by combined OTA and CIT.

DNA Damage Levels and DNA Repair Capability of Bone Marrow‐Derived Mesenchymal Stem Cells from Sprague Dawley® SPF rats as a Quality Parameter Before Surgical Transfection for the Treatment of Gestational Diabetic Myopathy

Mesenchymal stem cells (MSCs) have an important role for regenerative medicine through their self‐renewal and differentiation capability; besides its involvement in tissue development, maintenance, and repair, being useful for skeletal muscle regeneration by the production of secretory factors with trophic and engraftment functions. We propose to use the alkaline comet assay to evaluate the DNA damage levels and DNA repair capability according to cell culture passages as one of the parameters to test the quality of MSCs before surgical transplantation to the periurethral and rectus abdominis muscle (RAM) for the treatment of gestational diabetic myopathy in female rats at postpartum. MSCs were collected from bone marrow of female Sprague Dawley® rats. The characterization of these cells was done according to the International Society for Stem Cell. After isolation using Ficoll‐Paque and washing in low glucose Dulbecco's Modified Eagle Medium (DMEM) enriched with L‐glutamine, the cells were seeded in 25 cm2 plastic flasks containing high glucose DMEM enriched with L‐glutamine plus 20% bovine fetal serum, penicillin, streptomycin and amphotericin B. The cells were harvested at 90% of confluence and subjected to comet assay analysis. The slides were incubated in alkaline electrophoresis solution for 20 minutes for DNA unwinding and the run was conducted at 25v, 0.3A and 8W for 30 minutes, at 4 °C. The slides were stained with SYBR GOLD® (1:10.000 dilution) and analyzed in a fluorescent microscope using COMET ASSAY VI software. The DNA damage levels were ranked according to the percentage of damage as follows: 1) 0 % – 5% of damage; 2) 6% – 20% of damage; 3) 21% – 40% of damage; 4) 41% –95% of damage; 5) &gt; 95% of damage. The percentage of nucleoids with such levels of damage was calculated and analyzed using the Chi‐Square test. The analysis was conducted using SPSS (21.0) and p &lt; 0.05 was considered significant. This study was approved by the local Animal Ethics Committee (1234/2017 – CEUA). The partial results demonstrated that the 5th passage has a high percentage (70.04%) of cells with high levels of DNA damage (41% – 95%) assessed by tail intensity. The proportion of cells with high percentage of DNA damage (41% – 95%) is statistically significant from the proportion of cells with low DNA damage levels (0% – 5%, 6% –20%, and 21% –40%; p &lt; 0.001). The preliminary results demonstrated that the 5th passage is not useful for surgical transplantation because of high DNA damage levels. Future experiments will be conducted to test the DNA damage of the other passages and also evaluated the effect of cell culture on DNA repair capability. Furthermore, such results will be compared with the functional evaluations of periurethral and RAM after the surgical transplantation of MSC.Support or Funding InformationSupported by São Paulo Research Foundation (2016/01743‐5)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Alkaline comet assay results on fresh and one-year frozen whole blood in small volume without cryo-protection in a group of people with different health status

Using alkaline comet assay, DNA damage tail length (TL) and tail intensity (TI) parameters were compared between fresh whole blood and 1-year frozen small volume whole blood in EDTA at -80 °C without cryo-preservation. The studied group consisted of 25 volunteers with different health conditions who served as their own controls for frozen blood results. Without the purification step after thawing, the results and the usefulness of this protocol for future/retrospective (including re-analysations of putative outliers) studies were analysed. Medical surveillance and blood sampling were done at Merkur University Hospital Zagreb. No significant differences between fresh and frozen blood samples in terms of the mean TL values (mean ± SD: 29.03 ± 12.26 vs. 25.36 ± 6.97, respectively) and the mean TI values (9.19 ± 10.37 vs. 10.17 ± 8.55, respectively), and highly damaged cell percentage were determined among 25 volunteers. Median TI frozen samples values of entire group were within the allowed 10–11% (8.24). At the individual levels, no correlation between fresh and frozen whole blood samples was observed in 11 volunteers who suffered from diabetes mellitus type 2. Strong correlation between fresh/frozen samples was seen for TL (r = 0.64, p < 0.015) and TI (r = 0.71, p < 0.005) in nondiabetic subgroup. Overall, the results demonstrated the usefulness of the 1-year frozen blood without induction of heavily damaged DNA. Due to the different DNA damage behaviour connected with different health conditions, future studies should involve more volunteers, oxidative DNA damage comet assay measurements, the inclusion of a washing step after thawing and inclusion of disease/antioxidant biomarkers.

Aflatoxin production and in vitro toxicity of Aspergilli section Flavi isolated from air samples collected from different environments.

Aspergilli section Flavi, originally isolated from air samples collected from inhabited apartments (AP), unoccupied basements (BS), and processing facilities of a grain mill (GM), were analyzed for their potential to produce aflatoxin B1 (AFB1) on solid media. The isolates were further characterized with regard to their cytotoxic, genotoxic, and pro-inflammatory properties in vitro. Aspergilli were identified based on partial calmodulin (CaM) gene sequencing; the producing capacities of isolates were analyzed by HPLC/FLD and confirmed by genes in biosynthesis (aflR, norA, omtA). In the grain mill, the Aspergilli section Flavi (up to 1.3 × 106cfu/m3) dominated by AFB1-producing Aspergillus flavus (71%, 4.5-5254ng/ml) which showed a serious health risk for workers. Living environments were not relevant sources of exposure. After 24h, AFB1 (1-100μmol/l) reduced cell viability (MTT test) in both A549 cells and THP-1 macrophage-like cells without reaching IC50. In A549 cells, the extract of the AFB1-producing A. flavus significantly decreased cell viability but not below 50%. THP-1 macrophage-like cells were more sensitive to both extracts, but IC50 was obtained only for the AFB1-producing strain (0.37mg/ml; AFB1 2.78μmol/l). AFB1 (1 and 10μmol/l) induced significant DNA damage (tail intensity, alkaline comet assay) in A549 cells in contrast to Aspergilli extracts. AFB1 elevated IL-6 and IL-8, while Aspergilli extracts increased IL-1β, TNF-α, and IL-17 release in THP-1 macrophages (ELISA). Chronic exposure to AFB1 and/or other metabolites in airborne A. flavus from occupational environments may stimulate epithelial damage of airways accompanied by lowered macrophage viability.

Genotoxic evaluation and plasma oxidative stress markers in copper smelters

Smelters are at elevated risk of oxidative stress and DNA damage because of metal fumes exposure. This study aimed to evaluate oxidative stress (antioxidant capacity, plasma malondialdehyde (MDA)) and DNA damage in 47 smelters and to compare the results with 33 non-exposed workers. The DNA damage was significantly higher in the exposed compared to the non-exposed group (p < .001), with the largest effect size for tail intensity. There was no significant difference in the MDA and total antioxidant capacity between exposed and non-exposed participants. Exposure to metal fumes could increase the primary DNA damage and accelerate the cellular aging process in smelters.

Investigation of spatial and temporal variation of particulate matter in vitro genotoxicity and cytotoxicity in relation to the elemental composition

Even though the outdoor air pollution and its major component Particulate Matter (PM) are recently classified as human carcinogen, attempts to elucidate the underlying mechanisms of PM toxicity are still crucial and continuing with in vitro approaches in various environmental circumstances. Present study investigated the genotoxicity (Comet assay) and the cytotoxicity (lactate dehydrogenase (LDH) leakage and the water-soluble tetrazolium (WST-1) assays) of 30 daily PM2.5 samples collected in the Kütahya province, to address their daily variability in effects with season (i.e. winter versus summer) and location (i.e. rural versus urban) using A549 human lung cancer epithelial cell line, as well as in relation to their chemical composition, specifically trace elements, organic carbon (OC) and elemental carbon (EC). The genotoxicity, measured by the percentage tail intensity (TI), of the daily PM2.5 samples at the traffic dense urban station was higher than that of the rural site for 80% of the parallel days. The genotoxicity was significant in the winter at the urban and in the summer at the rural site. Cytotoxicity was the highest for the winter urban samples. The PM2.5 mass, OC, and EC concentrations were not correlated to DNA damage, while there were correlations with Mn, Fe, Cu and Ba at the rural PM2.5 samples, and Mn, Co and Ni at the urban samples, respectively. The present study is confirming that the complex composition of PM2.5 originating from spatial and temporal changes can cause differences in the health effects.

Single-strand DNA breaks and oxidative changes in rat testes exposed to radiofrequency radiation emitted from cellular phones

The testes are a sensitive organ to electromagnetic pollution and people are concerned about the harmful effects of the radiofrequency radiation (RFR) emitted from cellular phones. Therefore, the purpose of this study was to investigate the effects of long-term exposure to different RFR frequencies on single-strand DNA breaks and oxidative changes in rat testicular tissue. Twenty-eight male Sprague–Dawley rats were divided randomly into four groups. Three groups were exposed to radiation emitted from 900, 1800 and 2100 MHz RF generators, 2 h/day for 6 months. The sham-control group was kept under the same experimental conditions but the RFR generator was turned off. Immediately after the last exposure, testes were removed and DNA damage, 8-hydroxydeoxyguanosine (8-OHdG), malondialdehyde (MDA), total antioxidant status (TAS), total oxidant status (TOS) and oxidative stress index (OSI) were analyzed. The results of this study indicated that RFR increased TOS, OSI, MDA and 8-OHdG (p < 0.05). TAS levels in the exposed group were lower than in the sham group (p < 0.05). In terms of DNA damage, the tail intensities in the comet assay were higher in the exposure groups (p < 0.05). This study demonstrated that long-term exposure to RFR emitted by cellular phones may cause oxidative stress and oxidative DNA damage in rat testicular tissue and may generate DNA single-strand breaks at high frequencies (1800 and 2100 MHz). Our results showed that some RFR emitted from cellular phones has potential to lead to cell damage in the testes.

Comet assay and cytogenetic findings in differential diagnosis of Fanconi anemia

Fanconi anemia (FA) is a complex genetic disease with a variety of congenital and hematological symptoms, including the predisposition for cancer development. The main hallmark of FA cells, an increased chromosomal fragility, in the presence of the DNA-interstrand cross-linking chemicals, mitomycin C or diepoxybutane (DEB), makes the diagnosis of FA much easier. Cytogenetic method can detect the FA patients with highly elevated chromosomal breakage, but also some of the patients with borderline sensitivity to DEB no matter if they have FA or not. These particular circumstances lead us to introduce comet assay along with cytogenetic analysis, in order to determine DNA lesions and chromosomal fragility in untreated and DEB-treated lymphocytes of full blood from seven patients with clinical features of FA. Highly elevated DEB induced chromosomal sensitivity confirmed the diagnosis in five patients (FA group: 0.48-4.47 breaks/cell vs control group: 0.00-0.08 breaks/cell). Borderline DEB sensitivity (FA* group: 0.15-0.44 breaks/cell) was found in the remaining two patients. Results of the comet assay showed higher baseline and DEB-induced DNA damage values (Olive tail moment and tail intensity) in all five FA and one FA

Diffraction line profiles of spherical hollow nanocrystals

An analytical expression of diffraction line profiles of spherical hollow nanocrystals (NCs) is derived. The particular features of the profile lines, enhanced peak tail intensity, are analyzed and discussed as a function of the NC size parameters (outer and inner radius, shell thickness). The explicit formula for the integral breadth, the Fourier particle size, and the Scherrer constants are also obtained and discussed in detail. The diffraction line profiles of hollow CdS NCs of zincblende and wurtzite crystallographic structure are calculated and compared with Debye scattering profiles. The diffraction profiles of both approaches exhibit an enhanced peak tail intensity that can be considered as a fingerprint of the hollow NC structure.

Evaluation of the DNA damage in lymphocytes, sperm and buccal cells of workers under environmental and occupational boron exposure conditions

Industrial production and use of boron compounds have increased during the last decades, especially for the manufacture of borosilicate glass, fiberglass, metal alloys and flame retardants. This study was conducted in two districts of Balıkesir; Bandırma and Bigadic, which geographically belong to the Marmara Region of Turkey. Bandırma is the production and exportation zone for the produced boric acid and some borates and Bigadic has the largest B deposits in Turkey. 102 male workers who were occupationally exposed to boron from Bandırma and 110 workers who were occupationally and environmentally exposed to boron from Bigadic participated to our study. In this study the DNA damage in the sperm, blood and buccal cells of 212 males was evaluated by comet and micronucleus assays. No significant increase in the DNA damage in blood, sperm and buccal cells was observed in the residents exposed to boron both occupationally and environmentally (p = 0.861) for Comet test in the sperm samples, p = 0.116 for Comet test in the lymphocyte samples, p = 0.042 for micronucleus (MN) test, p = 0.955 for binucleated cells (BN), p = 1.486 for condensed chromatin (CC), p = 0.455 for karyorrhectic cells (KHC), p = 0.541 for karyolitic cells (KLY), p = 1.057 for pyknotic cells (PHC), p = 0.331 for nuclear bud (NBUD)). No correlations were seen between blood boron levels and tail intensity values of the sperm samples, lymphocyte samples, frequencies of MN, BN, KHC, KYL, PHC and NBUD. The results of this study came to the same conclusions of the previous studies that boron does not induce DNA damage even under extreme exposure conditions.

Optimization of the harvesting and freezing conditions of human cell lines for DNA damage analysis by the alkaline comet assay

The comet assay is a commonly used method for in vitro and in vivo genotoxicity assessment. This versatile assay can be performed in a wide range of tissues and cell types. Although most of the studies use samples immediately processed after collection, frozen biological samples can also be used. The present study aimed to optimize a collection and freezing protocol to minimize the DNA damage associated with these procedures in human cell line samples for comet assay analysis. This study was conducted in glial A172 and lung alveolar epithelial A549 cells. Two cell detachment methods (mechanical vs enzymatic) and two cryoprotective media [FBS + 10% DMSO vs Cell Culture Media (CCM) + 10% DMSO] were tested, and DNA damage assessed at four time points following storage at −80 °C (one, two, four and eight weeks). In both cell lines, no differences in % tail intensity were detected between fresh and frozen cells up to eight weeks, irrespective of the harvesting method and freezing medium used. However, freshly isolated A172 cells exhibited a significant lower DNA damage when resuspended in CCM + 10% DMSO, while for A549 fresh cells the preferable harvesting method was the enzymatic one since it induced less DNA damage. Although both harvesting methods and cryoprotective media tested were found suitable, our data indicate that enzymatic harvesting and cryopreservation in CCM + 10% DMSO is a preferable method for DNA integrity preservation of human cell line samples for comet assay analysis. Our data also suggest that CCM is a preferable and cost-effective alternative to FBS in cryopreservation media. This optimized protocol allows the analysis of in vitro cell samples collected and frozen at different locations, with minimal interference on the basal DNA strand break levels in samples kept frozen up to eight weeks.