Sister Chromatid Exchange Analysis Research Articles

Biosynthesis, characterization, toxicity assessment and bio-efficacy of silver nanoparticles synthesized by Microbacterium mitrae in controlling early blight in tomato (Lycopersicon esculentum L.)

This study evaluated the biogenesis of silver nanoparticles (NPs) using Microbacterium mitrae and its possibilities against phytopathogens. Silver nanoparticles have been synthesized using a simple protocol and the synthesized NPs have been characterized using UV visible spectroscopy, particle size analyzer, transmission electron microscope (TEM). Small-sized, stable silver nanoparticles in the range of 10-25 nm have been reported. Their antimicrobial activity against three phytopathogens i.e. Alternaria solani, Thanatephorus cucumeris and Botryodiplodia theobromae, was studied. The nanoparticles showed effective antimicrobial activity against all the tested fungal pathogens under in vitro conditions. The genotoxicity of NPs was also studied using sister chromatid exchange analysis on human lymphocytes and no adverse effects were observed up to the concentration of 100μg/ml. Efficient control against early blight has been observed within cytotoxicity limits in pot study, preventing the disease outbreak in tomato plants.

Mutagenic effect of the sex reversal hormone Diethylstilbestrol in the Nile tilapia (Oreochromis niloticus, L) through sister chromatid exchange analysis

The current study was conducted to explore the mutagenic effect of DES hormone on the treated Nile tilapia chromosomes using sister chromatid exchange (SCE) analysis. Two concentrations of DES hormone (100mg DES/kg and 200mg DES/kg diet) were addressed. The fry were fed for 60 days at 15% of the body weight daily. Two cell replication cycles, in the presence of 5-bromodeoxyuridine (BrdU) were achieved, and intestine tissues were used for chromosomes preparation. For slide staining, acridine orange (AO) was added, and the fluorescent microscope was used to the observation of AO fluorescence of stained slides. The obtained results indicated that the frequency of SCEs per cell and the percentage of cells exhibiting three SCEs per cell increased with the increasing hormone concentration. The present findings confirmed the mutagenic effect of DES hormone and clarified the possibility of using O. niloticus as an in vivo system for detecting mutagenic and carcinogenic chemicals. In addition, it is recommended not to use the hormone directly in the food served to humans. However, it can be used in the induction of fish sex inversion to XY females and followed by progeny testing, to generate super male YY.

Gene expression profile suggesting immunological dysregulation in two Brazilian Bloom's syndrome cases.

BackgroundBloom syndrome (BS) is a rare autosomal recessive chromosome instability disorder. The main clinical manifestations are growth deficiency, telangiectasic facial erythema, immunodeficiency, and increased risk to develop neoplasias at early age. Cytogenetic test for sister chromatid exchanges (SCEs) is used as a diagnostic marker for BS. In addition, most patients also present mutations in the BLM gene, related to defects in the DNA repair mechanism. However, the molecular mechanism behind the pathogenicity of BS is still not completely understood.MethodsWe describe two patients confirmed with BS by SCE and molecular analysis. Also, we performed the gene expression profile by the RNA‐seq methodology in mRNA transcripts for differential gene expression analysis using as a biological condition for comparison BS versus health controls.ResultsWe detected 216 differentially expressed genes related to immunological pathways such as positive regulation and activation of B cells, immune effector process and absence of difference of DNA repair genes expression. In addition; we also observed differentially expressed genes associated with apoptosis control, such as BCL2L1, CASP7, CDKN1A, E2F2, ITPR, CD274, TNFAIP6, TNFRSF25, TNFRSF13C, and TNFRSF17.ConclusionOur results suggest that the combination of altered expression of genes involved in signaling pathways of immune response and apoptosis control may contribute directly to the main characteristics observed in BS, such as recurrent infections, growth failure, and high risk of cancer. Transcriptome studies of other instability syndromes could allow a more accurate analysis of the relevant gene interactions associated with the destabilization of the genome. This is a first description of the profile of differential gene expression related to immunological aspects detected in patients with BS by RNA‐seq.

Genotoxicity test methods - a tool for DNA and chromosome damage biomonitoring

Nowadays, life is highly influenced by intense growth of various industries, high levels of pollution, and other environmental factors with harmful effects on human health. Therefore, cytogenetic monitoring is essential for detection of changes in the structure of chromosomes, which occur because of the effects of various genotoxic agents. In this review, we shall apprize the theoretical and experimental aspects of several tests for assessment of genotoxicity in humans such as Micronucleus assay, Comet assay, Chromosomal aberrations assessment and Sister chromatid exchanges analysis. These methods are accepted by the World Health Organization as standard tests for genotoxicological screening in humans. The methods are sensitive and confirm the cellular genotoxic effects of various genotoxicants.

4-Vinylbenzyl and 2-morpholinoethyl substituted ruthenium (II) complexes: Design, synthesis, and biological evaluation

Abstract Recently, the medical applications of organoruthenium complexes have attention attracted to organometallic chemists and biochemists. In this study, 4-vinylbenzyl and 2-morpholinoethyl substituted (NHC)Ru(II)(η6-p-cymene) complexes were synthesized and in vitro DNA binding, cell cytotoxicity (anticancer activity) and genotoxicity properties were determined in order to understand the biological activities. These complexes have been prepared from the 4-vinylbenzyl and 2-morpholinoethyl substituted Ag(I)NHC complexes via transmetallation method. The characterization of the new (NHC)Ru(II)(η6-p-cymene) complexes was performed by using 1H NMR, 13C NMR, FTIR spectroscopy and elemental analysis techniques. Also, molecular structure of complex 1b was obtained with single-crystal X-ray diffraction method. IC50 value of 1b against MCF-7 cell line was determined as 3.61 μM and for 1b, an oxidation effect was observed in the concentrations higher than 50 μg/mL. Comet assay unlike sister chromatid exchange (SCE) analysis data showed correlated results with cytotoxicity as well as DNA binding properties of the complexes.

Increased Non-Homologous End Joining Makes DNA-PK a Promising Target for Therapeutic Intervention in Uveal Melanoma.

Uveal melanoma (UM) is the most common primary intraocular tumour in adults, with a mean survival of six months following metastasis. The survival rates have not improved in over 30 years. This study has shown that sister chromatid exchange (SCE) is low in UM which is likely due to a reduced expression of FANCD2. As FANCD2 can function to suppress non-homologous end joining (NHEJ), this study therefore investigated NHEJ in UM. The activation of the catalytic subunit of the NHEJ pathway protein DNA-dependent protein kinase (DNA-PK) was measured by analysing the foci formation and the ligation efficiency by NHEJ determined using a plasmid-based end-joining assay. Using small-interfering RNA (siRNA) knock-down, and chemical inhibitors of DNA-PK, the survival of primary UM cultures and two cell lines were determined. To assess the homologous recombination capacity in response to the inhibition of DNA-PK, a SCE analysis was performed. In addition, to support the findings, the messenger RNA (mRNA) expression of genes associated with NHEJ was analysed using the Cancer Genome Atlas (TCGA)-UM RNAseq data (n = 79). The NHEJ activity and DNA-PKcs activation was upregulated in UM and the inhibition of DNA-PK selectively induced apoptosis and sensitized to ionising radiation and inter-strand cross-linking agents. The inhibition of the NHEJ protein DNA-PK is lethal to UM, indicating a potentially effective therapeutic option, either alone or as a sensitizer for other treatments.

Analysis of Sister Chromatid Exchanges and Proliferation of Human Peripheral Blood Lymphocytes Exposed to Epoxiconazole

Abstract The potential genotoxic/cytotoxic effect of epoxiconazole was evaluated by means of sister chromatid exchanges (SCE) following the 24 and 48 h in vitro exposure of human peripheral blood lymphocytes to epoxiconazole at concentrations of: 5, 10, 25, 50 and 100 μg. ml–1. Dimethyl sulphoxide (DMSO), used as an epoxiconazole solvent, was used as a negative control and mitomycine (MMC) as a positive control. After the 24-hour exposure, we failed to observe a significant increase in SCE frequencies in comparison with the negative control, however, the concentrations of 10—100 μg.ml–1 caused a significant decrease in the proliferation index (PI; P < 0.001). Also, the 48-hour exposure produced no significant alterations in the SCE frequencies in comparison with the control. At epoxiconazole concentrations ranging from 10 to 50 μg.ml–1 we recorded a moderate to strong, dose-dependent inhibition of PI (P < 0.05; P < 0.01; P < 0.001), while at the highest dose (100 μg.ml–1) the reduction in PI compared to the control was less pronounced (P < 0.05). The reduction in PI at the concentration range of 10—100 μg.ml–1 depended on the number of cells in the M1, M2 and M3 phases of the cell cycle per total number of 100 evaluated metaphases. Our results indicated a significant cytotoxic or cytostatic effect on human peripheral blood lymphocytes.

Effect of Methotrexate and Omega-3 Combination on Cytogenetic Changes of Bone Marrow and Some Enzymatic Antioxidants: An Experimental Study

Effect of Methotrexate and Omega-3 Combination on Cytogenetic Changes of Bone Marrow and Some Enzymatic Antioxidants: An Experimental Study

Effect of Methotrexate and Omega-3 Combination on Cytogenetic Changes of Bone Marrow and Some Enzymatic Antioxidants: An Experimental Study

Effect of Methotrexate and Omega-3 Combination on Cytogenetic Changes of Bone Marrow and Some Enzymatic Antioxidants: An Experimental Study

The evaluation of long-term effects of ionizing radiation through measurement of current sister chromatid exchange (SCE) rates in radiology technologists, compared with previous SCE values

Ionizing radiation is a strong physical mutagen, causing breakage of phosphodiester bonds in DNA at any stage of the mitotic cycle. Analysis of sister chromatid exchange (SCE) has come into use as a sensitive DNA-damage indicator. We investigated the SCE rates in radiology technologists who are occupationally and chronically exposed to ionizing radiation. The study included 39 radiology technologists and 35 sex- and age-matched healthy controls.There was a statistically significant difference in the SCE frequency between radiology technologists and controls (p<0.0001). Additionally, previous SCE data of 10 radiology technologists were compared with current results regarding radiation exposure time. There was statistically significant difference between previous and current SCE values (p=0.005).The significant increase in the frequency of SCE in radiology technologists emphasizes the importance of radiation–protection procedures in order to minimize radiation exposure and avoid possible genotoxic effects. Comparison of two studies that measured SCE values of radiology technologists after 8 years also suggests that the genotoxic effect is reversible.In conclusion, radiation is still an important mutagenic agent despite improvements in daily working hours and conditions.

Assessment of potential in vitro genotoxic and cytotoxic effects of bupropion hydrochloride (wellbutrin) in human peripheral lymphocytes and human cortical neuron

Introduction:Wellbutrin (bupropion hydrochloride; WB), an anti-depressant of the aminoketone class is new highly selective norepinephrine and dopamine reuptake inhibitor; it is effective in the treatment of patients with major depression.Materials and Methods:To investigate the in vitro effects of WB in human cultured peripheral blood lymphocytes and human cortical neural (HCN2) cell lines, micronucleus, sister chromatid exchange analysis, cellular viability, and comet assays were employed. The present study is to our knowledge, the first report on WB genotoxicity in cultured human peripheral blood lymphocytes and its cytotoxicity in the HCN2 cell line. We have also investigated the genotoxic potential of WB to induce chromosomal aberrations.Results:WB-induced cytotoxicity (measured as reduction of the nuclear division index) possibly prevented the division of damaged cells.Conclusion:We conclude that although, WB exerts potential genotoxic effects in cultured lymphocytes, its cytogenetic effects are very unlikely to occur in blood cultures of WB-administered subjects.

Cytogenetic analyses of Azadirachtin reveal absence of genotoxicity but marked antiproliferative effects in human lymphocytes and CHO cells in vitro

In this work we have examined the genotoxic potential of the bioinsecticide Azadirachtin A (AZA) and its influence on cell proliferation on human lymphocytes and Chinese Hamster ovary (CHO) cells. AZA genotoxicity was assessed by the analysis of chromosomal aberrations and sister chromatid exchanges (SCEs) in the absence and presence of rat liver S9 metabolism. Primary DNA damage was also investigated by means of the comet assay. The results obtained clearly indicate that AZA is not genotoxic in mammalian cells. On the other hand, AZA proved to interfere with cell cycle progression as shown by modulation of frequencies of first (M1) and second division (M2) metaphases detected by 5-Bromo-2′-deoxyuridine labeling. Accumulation of M1 metaphases were more pronounced in human lymphocytes. In the transformed CHO cell line, however, significant increases of multinucleated interphases and polyploid cells were observed at long treatment time. At higher dose-levels, the incidence of polyploidy was close to 100%. Identification of spindle structure and number of centrosomes by fluorescent immunostaining with α- and γ-tubulin antibodies revealed aberrant mitoses exhibiting multipolar spindles with several centrosomal signals.These findings suggest that AZA can act either through a stabilizing activity of microtubules or by inhibition of Aurora A, since both mechanisms are able to generate genetically unstable polyploid cells with multipolar spindles and multinucleated interphases.

Sister chromatid exchange analysis in some Holstein bulls

Summary Chromosomal appearance of 12 Holstein bulls selected for artificial inseminations were examined by sister chromatid exchanges (SCEs). For differential staining of sister chromatids bromodeoxyuridine (BrdU) was inoculated in lymphocyte cultures. The mean, maximum and minimum number of SCE per cell were determined 6.8 ± 1.14, 8.3 ± 1.1 and 5.7 ± 1.5, respectively. SCE frequencies of all animals were in the normal level. Also, evaluation of different breeding condition and age range did not show any significant statistical (P>0.05) effect on SCEs rates.

Role of oxidative stress in preeclampsia and intrauterine growth restriction

The aim of the present study was to evaluate the role of oxidative stress and DNA damage in preeclampsia and intrauterine growth restriction (IUGR). Twenty-four patients with preeclampsia, 20 patients with IUGR fetus and 37 healthy pregnant women were enrolled in the study. The total oxidant status (TOS) and antioxidant status (TAS) of plasma were measured using a novel automated colorimetric measurement method. Sister chromatid exchange (SCE) and micronuclei analysis were performed on peripheral blood lymphocytes of cases and controls. Women whose pregnancies were complicated with preeclampsia and IUGR had elevated levels of TOS and TAS when compared with healthy pregnant women (median TOS values: 9.73, 10.6 and 8.06, P = 0.001; median TAS values: 1. 77, 1.54 and 1.44, P < 0.001, respectively). The frequencies of SCE were only found to be increased in women with IUGR fetus compared with healthy pregnant women (8.81 vs 7.5, respectively, P = 0.02). Multivariable linear regression analysis for both TOS and TAS showed a significant relation between these variables and uric acid. Increased oxidative stress and antioxidative defense mechanisms may contribute to disease processes both in preeclampsia and IUGR.

Hydrocarbons and Kidney Damage: Potential Use of Neutrophil Gelatinase-Associated Lipocalin and Sister Chromatide Exchange

Background: Millions of workers are exposed to polycyclic aromatic hydrocarbons (PAHs) and it is known that the kidney is a target for toxic chemicals. We have evaluated neutrophil gelatinase-associated lipocalin (NGAL) as a potential marker of tubular damage and have used it, with sister chromatid exchange (SCE) analysis, to evaluate carcinogenic risk in a group of workers from an oil refinery. Methods: NGAL and SCE analysis were evaluated in 160 subjects. Exposed subjects were divided into three groups, according to levels of exposure to PAHs: 40 highly exposed workmen (WM), 40 less exposed office workers (OW), and 40 subjects (GE) living in Gela. The control group included 40 healthy subjects (HS). Results: WM, OW and GE showed higher NGAL levels than HS. WM had higher levels of NGAL than the OW and GE groups; in ROC analysis, serum NGAL showed a good diagnostic profile (sensitivity 87.5%; specificity 100.0%), as did urinary NGAL (sensitivity 90.0%; specificity 92.5%). Moreover, regarding SCE analysis, WM showed higher values than HS. A direct correlation between SCE and serum NGAL was found in WM, the group most exposed to PAHs. Conclusion: The high values of NGAL are an expression of damage to the renal tubule determined by exposure to PAHs. Compared to the other groups studied, chromosomal aberrations – expressed as SCE – were increased in WM, the group most exposed to PAHs, indicating genotoxic damage. NGAL may also play a role in the process of carcinogenesis having a direct correlation with the number of SCEs.

P53 null Fluorescent Yellow Direct Repeat (FYDR) mice have normal levels of homologous recombination

The tumor suppressor p53 is a transcription factor whose function is critical for maintaining genomic stability in mammalian cells. In response to DNA damage, p53 initiates a signaling cascade that results in cell cycle arrest, DNA repair or, if the damage is severe, programmed cell death. In addition, p53 interacts with repair proteins involved in homologous recombination. Mitotic homologous recombination (HR) plays an essential role in the repair of double-strand breaks (DSBs) and broken replication forks. Loss of function of either p53 or HR leads to an increased risk of cancer. Given the importance of both p53 and HR in maintaining genomic integrity, we analyzed the effect of p53 on HR in vivo using Fluorescent Yellow Direct Repeat (FYDR) mice as well as with the sister chromatid exchange (SCE) assay. FYDR mice carry a direct repeat substrate in which an HR event can yield a fluorescent phenotype. Here, we show that p53 status does not significantly affect spontaneous HR in adult pancreatic cells in vivo or in primary fibroblasts in vitro when assessed using the FYDR substrate and SCEs. In addition, primary fibroblasts from p53 null mice do not show increased susceptibility to DNA damage-induced HR when challenged with mitomycin C. Taken together, the FYDR assay and SCE analysis indicate that, for some tissues and cell types, p53 status does not greatly impact HR.

Genotoxic hazard evaluation in welders occupationally exposed to extremely low-frequency magnetic fields (ELF-MF)

Electric arc welding is known to involve considerable exposure to extremely low-frequency magnetic fields (ELF-MF). A cytogenetic monitoring study was carried out in a group of welders to investigate the genotoxic risk of occupational exposure to ELF-MF. This study assessed individual occupational exposure to ELF-MF using a personal magnetic-field dosimeter, and the cytogenetic effects were examined by comparing micronuclei (MN) and sister chromatid exchange (SCE) frequencies in the lymphocytes of the exposed workers with those of non-exposed control subjects (blood donors) matched for age and smoking habit. Cytogenetic analyses were carried out on 21 workers enrolled from two different welding companies in Central Italy and compared to 21 controls. Some differences between the groups were observed on analysis of SCE and MN, whereas replication indices in the exposed were found not to differ from the controls. In particular, the exposed group showed a significantly higher frequency of MN (group mean±SEM: 6.10±0.39) compared to the control group (4.45±0.30). Moreover, the increase in MN is associated with a proportional increase in ELF-MF exposure levels with a dose-response relationship. A significant decrease in SCE frequency was observed in exposed subjects (3.73±0.21) compared to controls (4.89±0.12). The hypothesis of a correlation between genotoxic assays and ELF-MF exposure value was partially supported, especially as regards MN assay. Since these results are derived from a small-scale pilot study, a larger scale study should be undertaken.

Preface

Preface

Genoprotective Effect of Indian Gentian in Type 2 Diabetes Mellitus (T2DM): Comet Assay, Sister Chromatid Exchange and Protein Oxidation Studies

The present study was undertaken to study the effect of Indian Gentian (Enicostemma littorale Blume), a herb, as a genoprotective agent in type 2 diabetes mellitus (T2DM) patients. For this, a total of 52 T2DM patients were investigated, of which 38 received 500 mg - 1 g Indian Gentian thrice daily escalated over three months (Group 1). The remaining 14 patients were not given the herb (Group 2). Fifteen age and sex matched non diabetic healthy volunteers served as controls (Group 3). All three groups were studied for DNA damage by comet assay and Sister Chromatid Exchanges (SCEs); Group 1 was also investigated for protein oxidation. Paired and unpaired t tests were performed at 95% confidence interval. Results of comet assay and SCE studies revealed that in Group 1, post Indian Gentian treatment, normal cell population increased, whereas moderately damaged, highly damaged and apoptotic cell population and SCE decreased as compared to Group 1 (pre-treatment patients) and Group 2 (without treatment patients). In comet assay , statistically significant dif ference between Group 1 (post-treatment patients) and Group 3 (controls) suggested that the herb was able to decrease the DNA damage but not as low as non-diabetic healthy controls. On the other hand, SCE analysis showed that the herb can reduce such exchanges to as low as the controls. In protein oxidation assay , no significant difference was found between the pre- and post-treatment T2DM patients of Group 1. The present study therefore indicated that overall Indian Gentian may have a significant ef fect on reducing DNA damage and attenuating SCEs in T2DM patients.

In vitro genotoxicity of rocuronium bromide in human peripheral lymphocytes

Rocuronium bromide (RB), an aminosteroid type neuromuscular blocking agent, acts by reducing or inhibiting the depolarising effect of acetylcholine on the terminal disc of the muscle cell. To our knowledge, there is no adequate information on the genotoxic effects of RB, up to now. In the present study, possible genotoxic effects of RB have been determined by means of sister chromatid exchange (SCE), chromosome aberration (CA) and micronucleus (MN) analyses in human peripheral blood lymphocytes. The human peripheral blood lymphocytes were exposed to three different concentrations of RB (60, 80 and 100μg/mL) for 24- and 48-h. In this study, RB increased the frequency of CAs, however, did not increase the frequency of SCEs. RB did not decrease the proliferation index (PI) and mitotic index (MI). Accordingly, RB increased the frequency of micronucleus (MN) but did not decrease the nuclear division index (NDI). Findings from this study suggest that rocuronium bromide is clastogenic but not cytotoxic to cultured human peripheral blood lymphocytes.