DNA Damage Index Research Articles

Mangifera indica L.: Assessing the cytogenotoxicity of aqueous extract through micronucleus and comet assays

Mangifera indica L., known as mango tree, is a species belonging to family Anacardiaceae, widely used in popular medicine whose leaves are used to treat several diseases. Although several studies did not detect the cytogenotoxic activity of M. indica leaf extract, other authors have observed these feature and highlighted the importance of further research about the toxic potential of M. indica leaf extract. The aim of the current research is to assess the cytogenotoxic potential of the aqueous extract of M. indica leaves through comet DNA assay and micronucleus test. It was possible observing significant increase in the number of micronucleated polychromatic erythrocytes (MNPCE) in mice subjected to the treatment with the extract, in addition to significant increase in DNA damage index in comparison to the negative control group. The extract also presented cytotoxic effect caused by significant reduction in the polychromatic:normochromatic erythrocyte (PCE/NCE) ratio in comparison to the negative control group. The extract of M. indica leaves is cytogenotoxic under the conditions used in the current research; this outcome highlighted the importance of comparative studies applied to the several used processing methods, as well as to cultivation conditions, to certify the safety in the popular use of medicinal plants.

Perftoran ® Inhibits Hypoxia-Associated Resistance in Lung Cancer Cells to Carboplatin.

Perftoran® (perfluorodecalin) is an oxygen carrier, and carboplatin is a common chemotherapy drug used worldwide for lung cancer treatment. Hypoxia is one of the factors that induce resistance of lung cancer cells to carboplatin. This study explored the role of Perftoran®, as an oxygen carrier, in lowering the resistance of lung cancer cells to carboplatin through suppression of hypoxia pathway mediators. The effect of Perftoran® on the resistance of human lung cancer A549 cells to carboplatin was investigated through the evaluation of cytotoxicity by MTT, cell death mode by dual DNA staining, DNA damage by comet assay, DNA platination (DNA/carboplatin adducts) by atomic absorption spectroscopy, hypoxia degree by pimonidazole, HIF-1α/HIF-2α concentrations by ELISA, expression of miRNAs (hypoxamiRs miR-210, miR-21, and miR-181a) by qRT-PCR, and the content of drug resistance transporter MRP-2 by immunocytochemical staining. Results indicated that compared to carboplatin, Perftoran®/carboplatin decreased cell resistance to carboplatin by potentiating its cytotoxicity using only 45% of carboplatin IC50 and inducing apoptosis. Perftoran® induced DNA platination and DNA damage index in cells compared to carboplatin alone. Moreover, compared to treatment with carboplatin alone, co-treatment of cells with Perftoran® and carboplatin inhibited cellular pimonidazole hypoxia adducts, diminished HIF-1α/HIF-2α concentrations, suppressed hypoxamiR expression, and decreased MRP-2. In conclusion, Perftoran® inhibited resistance of lung cancer cells to carboplatin through the inhibition of both hypoxia pathway mediators and the drug resistance transporter MRP-2 and through the induction of DNA/carboplatin adduct formation.

RETRACTED ARTICLE: Evaluating the role of propolis and bee venom on the oxidative stress induced by gamma rays in rats

Honeybee products consist of many substances, which have long been known for their medicinal and health-promoting properties. This study set out to appraise the protective potential of Egyptian propolis (EP) and bee venom (BV) separately or combined against total body irradiation (TBI) induced oxidative injury in rats. Besides, we assessed the bioactive components in EP and BV using HPLC and UPLC/ ESI–MS analysis in the positive ion mode. The animals were subjected to a source of gamma ionizing radiation at a dose of 6 Gy. Propolis and BV were administered independently and in combination before 14 days of γ-irradiation. Liver and kidney functions were estimated besides, DNA damage index (8- OHdG) by ELISA. Antioxidants, including glutathione (GSH), catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx) were detected. Gene expression technique investigated for BAX, BCL2, and in plasma also miR125b expression in serum of rats. Besides, the histopathological for the brain, liver, kidney, and heart were investigated. In addition, lipid peroxidation was investigated in plasma and in the previous organs. The present results provide opportunities to advance the use of bee products as promising medicinal sources.

Tadalafil and Sildenafil illicit association: Stability- indicating HPLC method, photodegradation kinetic and toxicological studies

The illicit market of counterfeit medicines containing sildenafil and tadalafil has been causing serious public health problems. Thus, further studies on this illicit association are needed. A stability-indicating HPLC method was developed for simultaneous determination of tadalafil (TAD) and sildenafil (SIL) using a C18 column (250 x 4.6 mm, 5 µm). Detection was achieved at 284 nm, for TAD, and 292 nm, for SIL. The method was considered to be specific, linear, precise, accurate, robust, and sensitive. In the photodegradation kinetic studies, the drugs showed a first-order reaction rate when isolated, and zero-order when associated. Toxicological assays demonstrated that the photodegraded drugs decreased cell viability in compared to non- degraded drugs, suggesting cytotoxic activity. Additional, mutagenic activity was not observed under the tested conditions. Photodegraded drugs, in association, depicted DNA damage index, suggesting genotoxic effects. The obtained results will be able to support the forensic intelligence laboratories, as well as to alert the population about the risk inherent to consuming counterfeit products.

Berberine increases the expression of cytokines and proteins linked to apoptosis in human melanoma cells

Melanoma is the most lethal form of skin cancer, and its incidence has increased considerably in the last decades. Melanoma presents difficult treatment with strong resistance of tumor cells, due to its extremely invasive nature with high capacity to metastases. Berberine (BBR), an isoquinoline alkaloid, is a molecule found in several medicinal plants, and has been studied in several diseases, demonstrating antimicrobial, antidiabetic and anti-inflammatory properties and anti-tumorigenic effects. In SK-MEL-28 cells, 50μM BBR treatment for 24h decreased cell viability by 50 percent. This concentration generated cell death both by early apoptosis and necrosis, with an increase in the DNA damage index. BBR increased (*p < 0.05) the proportion of cells in G1/G0 phase and decreased (###p < 0.005) the percentage of cells in S phase. The alcaloid increased (****p < 0.001) ROS production compared to untreated controls with an increase in activated caspase 3 and phosphorylated p53 protein levels. In addition, BBR significantly enhanced ERK as well as both pro- and anti-inflammatory cytokine expression compared to untreated controls. BBR has important antiproliferative effects and may be alone or in adjunct therapy a promising candidate for melanoma treatment, a cancer with great incidence and high lethality.

Short Term Effects of Antimicrobial Agent Triclosan on Oreochromis mossambicus (Peters, 1852): Biochemical and Genetic Alterations

Background: Triclosan is an antimicrobial agent which enters into the aquatic environment through wastewater discharges which causes potential health risk in human and aquatic organisms. The present study aimed to determine the toxic effects of triclosan on Oreochromis mossambicus. Methods: The fishes were subjected to five different concentrations viz. 131, 262, 523, 1046 and 2092 µg.l-1 of triclosan for 96 h acute toxicity test. To evaluate the levels of enzymes such as acetylcholinesterase and glutathione S transferase, brain and liver tissues were collected, homogenized, extracted and stored at -20° for further analysis. The DNA damage was assessed in gill and liver tissues using single cell gel electrophoresis method. Result: In present study, the calculated 96 h LC50 value of triclosan in O. mossambicus was 740 µg.l-1 and the fishes showed various behavioural alterations. Time and dose dependent inhibition of AChE activity in brain tissue was observed during acute toxicity test. However, the GST activity in liver tissue increased on exposure to triclosan with significant increase in concentration of toxicant. DNA damage index was higher in gill tissue compared to the liver tissue during acute exposure to TCS which could cause detrimental effects in fishes.

Toxicity effects of size fractions of incinerated sewage sludge bottom ash on human cell lines

Sewage sludge bottom ash (SSBA) from the incineration plant used for the production of construction materials possibly possess heavy metals which might cause a negative impact on human health. Considering biosafety, we investigated the toxicity effects of 0.5–2 mm (aggregate substitute) and < 0.075 mm (cement substitute) in its solid and leachate form on human lung fibroblast cells (MRC-5) and human skin epidermal cells (HaCaT) on exposure through contact. MTS assay revealed the cellular responses of lung and skin cell lines to the leachates showing that the skin cells, which often interact with the external environment displayed better tolerance than the lung cells, whereas solid ash showed a concentration and size-dependent toxicity. Solid ash was found to downregulate the intracellular glutathione/superoxide dismutase activities and upregulate lactate dehydrogenase/lipid peroxidation activities thus inducing oxidative stress to the cell and subsequently resulting in the cell membrane leakage, destructive mitochondrial membrane potential (Δψm), apoptosis, and DNA damage, which is nearly 7-fold higher than the negative control. At a high concentration, DNA damage index of 1.09 and 1.29 was observed for the 0.5–2 mm sized ash leachate on skin cells and lung cells respectively, whereas for ash (<0.075 mm size) leachate, this fraction was 1.29 and 2.96, respectively. Overall, the ash leachate is found to be safer/biocompatible if they come in contact with humans as compared to SSBA in its solid form.

Assessment of the cytoprotective effect of the homeopathic compound Canova® on African green monkey kidney (VERO) cell line exposed to the drug dipyrone sodium

ABSTRACT Dipyrone or metamizole is one of the most frequently used analgesic worldwide. Despite its widespread use, this drug may exert genotoxic and cytotoxic effects on lymphocytes. Therefore, studies with therapeutic agents that may provide protection against these effects are important. The homeopathic compound Canova® (CA) appears to be a beneficial candidate for preventing DNA damage and cellular lethality, since this compound acts as an immunomodulator associated with cytoprotective actions. Hence, the aim of the present investigation was to determine the potential cytoprotective effects of CA using cell line VERO as a model. VERO cells were incubated with sodium dipyrone and subsequently subject to the comet, apoptosis and immunocytochemistry assays. Data demonstrated that sodium dipyrone induced an increase in DNA damage index (DI) employing the comet assay. However, when VERO cells were co-treated with CA at the three concentrations studied, a significant reduction in DI was observed, indicating an antigenotoxic effect attributed to CA. Further dipyrone induced an elevation in %apoptosis at 24 and 48 hr. However, when dipyrone was co-incubated with CA, a significant reduction in %apoptosis was noted at the three concentrations of CA employed. Results from immunocytochemical analysis showed a rise in the expression of caspase 8 and cytochrome C when cells were exposed to dipyrone. In contrast, co-treatment of dipyrone and CA significantly reduced the effect of dipyrone. Therefore, evidence indicated that CA acted as an anticytotoxic and antigenotoxic agent counteracting damage induced by dipyrone.

TRESK Regulates Gm11874 to Induce Apoptosis of Spinal Cord Neurons via ATP5i Mediated Oxidative Stress and DNA Damage.

Reportedly, TWIK-related spinal cord K+ (TRESK) deficiency in spinal cord neurons positively correlates with the mechanism underlying neuropathic pain (NP). However, the precise effects of TRESK on neurons of the spinal cord remain elusive. In the present study, we investigated the impact of TRESK silencing on spinal cord neurons to further elucidate the downstream mechanisms of TRESK. Herein, neurons of the dorsal spinal cord were cultured as a cell model for investigations. Apoptosis, oxidative stress, and DNA damage-related proteins were evaluated. Additionally, flow cytometry, microarray profiling, real-time polymerase chain reaction (PCR), western blotting, fluorescence in situ hybridization (FISH), immunofluorescence, and enzyme-linked immunosorbent assay (ELISA) were performed. In cultured neurons, the downregulation of TRESK mRNA expression induced apoptosis of dorsal spinal cord neurons. Using real-time PCR and western blotting, the upregulation of LncRNA Gm11874 (Gm11874) and ATP5i, screened from the gene chip, was confirmed. On silencing TRESK, expression levels of γ-H2AX, poly [ADP-ribose] polymerase 1 (PARP-1), FoxO1, FoxO3, MitoSOX, malondialdehyde (MDA), and 8-hydroxy-2' -deoxyguanosine (8-OHdG), which are known indices of oxidative stress and DNA damage, were significantly elevated. Moreover, ATP induced oxidative stress, DNA damage, and apoptosis were reduced by ATP5i siRNA. Finally, Gm11874 and ATP5i were co-expressed in spinal cord neurons in a FISH experiment, and the expression of ATP5i was positively regulated by Gm11874. These results implied that ATP5i induced oxidative stress and DNA damage, resulting in neuronal apoptosis, and Gm11874 was confirmed to act upstream of ATP5i. Our study revealed that TRESK silencing upregulated Gm11874 to induce apoptosis of spinal cord neurons, which resulted in ATP5i promoting oxidative stress and DNA damage. These findings could highlight the TRESK-mediated NP mechanism.

Chronic Exposure to Toluene and Heavy Metals and Changes in Indices of Liver Function, Inflammation and Oxidative DNA Damage among Automobile Workers

Background: Oxidative stress (OS), oxidative DNA damage and inflammatory response induced by chronic exposure to volatile organic compounds and heavy metals (HM) have been implicated in multiple organ dysfunction. The liver enzymes (alanine aminotransferase (ALT), alkaline phosphatase (ALP), gamma glutamyl transferase (GGT)), biomarkers of OS (nitric oxide (NO), glutathione (GSH), total antioxidant capacity (TAC), total plasma peroxides (TPP), malondialdehyde (MDA)) oxidative stress index (OSI)), oxidative DNA damage (8-hydroxy-2-deoxyguanosine (8-OHdG)), and inflammation marker (tumor necrosis factor alpha (TNF-I±)); heavy metals (cadmium (Cd), lead (Pb)) and urine hippuric acid (uHA) levels were assessed in automobile workers. Methods: Fifty automobile workers and 50 controls aged 18-60 years were enrolled into this study. The MDA, GSH, NO, TAC, TPP, ALT, ALP and GGT were estimated by colorimetry, 8-OHdG and TNF-I± by enzyme linked immunosorbent assay, Cd, Pb by atomic absorption spectrophotometry and uHA by high performance liquid chromatography. Data were analyzed using t-test and correlation analysis at p <0.05. Results: Automobile workers had significantly higher liver enzymes, lipid peroxidation, oxidative stress, oxidative DNA damage, nitric oxide, HM, uHA and lower total antioxidants relative to controls. Heavy metals were positively associated with MDA, TPP and OSI; TPP with duration of exposure; ALP with number of working hours; and liver enzymes with OSI only in automobile workers. Conclusion: Association of exposure to toluene and heavy metals with increased liver enzymes activity, lipid peroxidation, oxidative stress, oxidative DNA damage, and depressed antioxidants in automobile workers suggest increased risk of hepatotoxicity and hepatocellular carcinogenesis.

Association between a marker of sperm DNA damage and sperm indices in infertile males in Benin City, Nigeria: A cross-sectional study.

Background Studies have shown oxidative DNA damage is associated with male infertility.ObjectiveThis study determines the levels of 8-hydroxy-2'-deoxyguanosine (8-OHdG) and some markers of oxidative stress in seminal fluid of males investigated for infertility and men of proven fertility in Benin City, Nigeria.Materials and MethodsSemen samples produced by self or assisted masturbation were analyzed by microscopic technique according to the World Health Organization guidelines. Thereafter, samples were centrifuged and seminal fluid plasma separated and stored at -20°C prior to assay for 8-OHdG and oxidative stress biomarkers. Based on the sperm concentration/count, the overall samples were grouped into the following categories: normospermia (n = 20), oligozoospermia (n = 30), and azoospermia (n = 20). The control group comprised of 30 age-matched males of proven fertility. The seminal fluid 8-OHdG, total antioxidant status, superoxide dismutase and malondialdehyde (MDA) were assayed through ELISA and spectrophotometric methods, respectively.ResultsSeminal plasma level of 8-OHdG and MDA were significantly higher (p = 0.01) in infertile subjects than controls. The mean levels of 8-OHdG and MDA in infertile subjects were higher in azoospermia than oligospermia than normospermia and so, was least in the normospermia. Conversely, the mean levels of total antioxidant status and superoxide dismutase were significantly lower (p = 0.01) in infertile than fertile the control male subjects with levels higher in normospermia than oligospermia and least in azoospermia. Moreover, the seminal 8-OHdG correlated negatively with sperm count (r = -0.359, p = 0.01), percent motility (r = -0.388, p = 0.04), and percent morphology (r = -0.327, p = 0.02).ConclusionThe assessment of sperm DNA damage in addition to routine seminal fluid analysis may play an important role in specific diagnosis and management of male infertility.

Detection of DNA Damage in the Frozen Chicken Meat from the Baghdad Local Markets Using the Single-cell Gel Electrophoresis Assay under Abusive Storage Conditions

Effects of abusive storage conditions were studied with the single-cell gel electrophoresis test by detecting DNA damage to the thighs and breasts (comet assay). Twenty frozen thigh and breast meat samples were taken from the local market of Baghdad city, and divided equally ten samples for each, and examined in triplicate in the experiment. The DNA damage index design, integrating the distribution of mean tail moments between two groups, in the present study took the DNA in head and tail parameters that provided values significantly different, which allowed more precise discrimination between samples according to the lousy condition storage levels. Breast cells were more sensitive to abusive conditions than thigh muscle cells. In conclusion, the DNA in the head was high significantly in the thigh meat samples more than breast meat samples, whereas the tail moment shows an increase in the breast meat significantly more than thigh meat samples.

Ku70 modulation alleviates murine allergic asthma features and restores mitochondrial function in lungs

The airway epithelium is continuously exposed to a variety of pollutants and allergens, thanks to both natural and manmade environmental pollution. With numerous protective mechanisms, the airway epithelium protects the lungs. DNA repair mechanism is one such protective response and its failure could lead to the accumulation of DNA mutations. Our lab had earlier demonstrated the dysfunctional mitochondria in airway epithelium of the asthmatic mice lungs. Here, we show that Ku70 modulation by the administration of Ku70 plasmid attenuates asthma features and reduces mitochondrial dysfunction in the lungs of allergen exposed mice. Ku70 is a key DNA repair protein with diverse roles including VDJ recombination, telomere maintenance, and maintenance of cell homeostasis. Recently, we found a reduction in Ku70 expression in asthmatic airway epithelium, and this was associated with mitochondrial dysfunction in asthmatic condition. In this study, we have shown that Ku70 over-expression in asthmatic mice attenuated airway hyperresponsiveness, airway inflammation, sub-epithelial fibrosis along with reduction in TGF-β with no effect in IL-13 levels and goblet cell metaplasia. Ku70 over-expression in asthmatic mice reduced 8-isoprostane, a marker of oxidative stress, and restored the mitochondrial function in asthmatic mice. We further found these roles of Ku70 to be independent of DNA damage as Ku70 overexpressed mice did not show any reduction in DNA tail, an index of DNA damage. Thus, our findings indicate that Ku70 can attenuate crucial features of asthma along with the restoration of mitochondrial function. This implies that Ku70 could be a therapeutic target for asthma without affecting DNA repair function.

Antioxidant, genotoxic, antigenotoxic, and antineoplastic activities of apitoxin produced by Apis mellifera in Northeast, Brazil

ABSTRACT: The objective was to evaluate the in vitro antioxidant, genotoxic, antigenotoxic, and antineoplastic activities of apitoxin produced by the bee Apis mellifera. The antioxidant activity of the apitoxin solution was evaluated using the DPPH (2,2-diphenyl-1-picrilhydrazyl) method. Genotoxic potential of apitoxin was analyzed by comparing the mean DNA damage indices (idDNA) of L929 strain fibroblasts exposed to hydrogen peroxide (H2O2 - genotoxic substance), distilled water, or apitoxin. The antigenotoxic effect of apitoxin was analyzed by assessing the percentage decrease in H2O2-induced genotoxicity in L929 fibroblasts co-treated with three concentrations of the aqueous apitoxin solution and subjected to comet assay. In vitro antineoplastic activity in human tumor cell lines of prostate adenocarcinoma (PC3), hepatocellular carcinoma (HEPGE2), melanoma (MAD-MB435), and astrocytoma (SNB19), were verified by MTT [3- (4) bromide colorimetric method, 5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium]. Apitoxin had no genotoxic effect on L929 cells at concentrations of 30, 10, and 5 µg/mL after 24 hours of exposure. This effect was only evident at 50 µg/mL. Apitoxin promoted a significant reduction in DNA damage index (idDNA) at all concentrations tested. At 30 µg/mL, apitoxin attenuated the genotoxic effects induced by H2O2. Apitoxin also demonstrated in vitro antineoplastic potential, since the cytotoxic effect was observed at concentrations of 50 µg/mL and 25 µg/mL, with significant reduction in viability percentage of PC3 tumor cell lines, HEPGE2, MAD-MB435, and SNB19. The high antioxidant activity associated with the absence of genotoxic effect and the genoprotective and antineoplastic effect demonstrated by apitoxin here provide indications of apitoxin’s therapeutic potential.

Evaluation of the Genotoxicity of Endodontic Materials for Deciduous Teeth Using the Comet Assay

Objective: To evaluate genotoxicity of zinc oxide, P. A. calcium hydroxide, mineral trioxide aggregate and an iodoform paste using comet assay on human lymphocytes. Material and Methods: Two positive controls were used: methyl-methanesulfonate for the P.A. calcium hydroxide and mineral trioxide aggregate; and doxorubicin for the iodoform paste and zinc oxide. There were also two negative controls: distilled water for the P.A. calcium hydroxide and mineral trioxide aggregate; and DMSO for the iodoform paste and zinc oxide. Comets were identified using fluorescence microscopy and 100 of them were counted on each of the three slides analyzed per drug test. A damage index was established, taking into consideration the score pattern that had previously been determined from the size and intensity of the comet tail. Analysis of variance, followed by Tukey’s test, was used to compare the means of the DNA damage indices. Results: The DNA damage index observed for mineral trioxide aggregate (7.08 to 8.58) and P.A. calcium hydroxide (6.50 to 8.33), which were similar to negative control index. On the other hand, damage index for zinc oxide (104.7 to 218.50) and iodoform paste (115.7 to 210.7) were similar to positive control index. Conclusion: Iodoform paste and zinc oxide showed genotoxicity at all concentrations used.

Effect of taurine associated gold nanoparticles on oxidative stress in muscle of mice exposed to overuse model.

Muscle overuse and its consequent muscle damage has no cure. Therefore, the present study aimed to investigate the regulatory role of tau-AuNPs on muscle recovery of muscle overuse model. The animals (Male Swiss mice) were randomly divided into four groups: Control (Ctr; n=6); tau-AuNPs (n=6); overuse (n=6); and overuse plus tau-AuNPs (n=6). Exercise sessions were performed for 21 consecutive days, and one exercise model was applied daily in the following sequence: low intensity, moderate intensity, and high intensity. The mice were then sacrificed. The quadriceps muscles were surgically removed for subsequent biochemical analysis (oxidative stress parameters, DNA damage markers and muscle differentiation protein). The overuse group significantly increased the oxidative stress parameters and DNA damage markers, whereas tau-AuNPs significantly decreased the oxidative stress parameters in the overuse animal model. However, there were no significant differences observed between overuse group and overuse plus tau-AuNPs administrated group in relation to DNA damage markers including DNA damage frequency and index levels when compared to control and tau-AuNPs groups. Muscle differentiation protein Myf-5 was increased in the overuse plus tau-AuNPs administration group when compared to control group. In conclusion, tau-AuNPs had significant effect on reducing oxidative stress parameters and increasing myogenic regulatory protein Myf-5 in the overuse group. However, it did not have significant effect on reducing DNA damage.

Geno-cytotoxicity and congenital malformations produced by relevant environmental concentrations of aluminum, diclofenac and their mixture on Cyprinus carpio. An interactions study

Several studies highlight the presence of aluminum and diclofenac in water bodies around the world and their ability to induce oxidative stress and a negative effect on biomolecules in several aquatic species. However, studies evaluating the toxic effect of mixtures of these contaminants are scarce. The objective of this work was to determine the genotoxic, cytotoxic and embryotoxic effect of the mixture of aluminum and diclofenac at environmentally relevant concentrations on Cyprinus carpio.Juveniles of Cyprinus carpio were exposed to 0.31 μg L−1 of diclofenac, 24.45 mg L-1 of aluminum, and a mixture of both contaminants at the same concentrations for 12, 24, 48, 72 and 96 h. After the exposure time the liver, gills and blood were extracted and the following biomarkers were evaluated: micronucleus frequency, comet assay, caspase activity and TUNEL test. On the other hand, Cyprinus carpio embryos were exposed to diclofenac (0.31 μg L−1), aluminum (0.06 mg L−1) and their mixture at the same concentrations and exposure time. Microscopic observation was performed to evaluate embryonic development at 12, 24, 48, 72 and 96 h.Diclofenac (0.31 μg L−1) induces significant increases in micronucleus frequency with respect to control (p < 0.05), in all tissues. Aluminum (24.45 mg L−1) significantly increases DNA damage index in liver and blood cells with respect to control (p < 0.05). All treatments increase caspases activity in all tissues with respect to control (p < 0.05). Diclofenac increases the percentage of TUNEL-positive cells in liver and blood; while aluminum and the mixture increases it significantly in gills and blood with respect to the control (p < 0.05). The mixture significantly delays embryonic development, while aluminum and the mixture significantly increase teratogenic index with respect to control (p < 0.05). In conclusion, exposure to environmental concentrations of aluminium, diclofenac and their mixture induces genotoxic damage, cell death by apoptosis and negative effects on the development of Cyprinus carpio and the toxic response is modified by the interaction of the xenobiotics.

Establishment of an in vitro model using the rat alveolar macrophage cell line NR8383.

To establish an in vitro model of radiation-induced lung injury using rat lung alveolar macrophages (NR8383). Using a medical electronic linear accelerator, cells were irradiated with either 0 Gy or 6 Gy X-rays. At 6, 12, 24, 30 and 48 h, the DNA damage index (8-OHdG) and lipid damage index (MDA) were measured in the two groups. We also determined the levels of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and transforming growth factor-β (TGF-β). The levels of 8-OHdG and MDA in the 6 Gy irradiation group were higher than those in the 0 Gy group at 6, 12, 24, 30 and 48 h after irradiation. The levels reached the highest value -6 h after irradiation, and then gradually decreased. The levels of the inflammatory factors TNF-α, TGF-β and IL-6 were higher in the 6 Gy irradiation group than those in the 0 Gy group at 6, 12, 24, 30 and 48 h after irradiation. Six Gy X-ray irradiated NR8383 cells can be used to establish an in-vitro model of radiation-induced lung injury. The levels of 8-OHdG, MDA, TNF-α, TGF-β and IL-6 can be used as effective evaluation indicators.

The effects of thermal and ethanolic stress in industrial strains of Saccharomyces cerevisiae

Saccharomyces cerevisiae is exceptional microorganisms used in biotechnological processes, mainly in the ethanol production chain. Studies of the cellular responses of industrial yeasts under ethanolic and thermal stress in an association are still incipient. This study aimed to evaluate the action of thermal and ethanolic stress in industrial strains of Saccharomyces cerevisiae under different temperatures and concentrations of ethanol, to understand whether these factors influence ethanol production. For cytotoxicity and genotoxicity tests, yeasts were grown in 2% YPD medium incubated for 10 hours at 250 rpm. After growth, the samples were grown in sugarcane juice in concentrations of 5, 10 and 15% ethanol and incubated at 30 and 40 ºC. In Petri dishes containing the solid medium YPD 2% the yeasts were dripped and incubated for 72 hours the cytotoxic action was analyzed by cell growth and genotoxicity through the comet assay and ethanol production by gas chromatography. Cell growth occurred in all conditions, however, at 30 ºC there was inhibition in 10% (v v-1) of ethanol being potentiated in 15% (v v-1), at 40 ºC. The genotoxicity analysis showed an induction of DNA damage in yeasts, however, the FLE yeast was the one with the highest DNA damage index. The yeast Pedra-2 was more tolerant and produced more ethanol, showing to be a tolerant strain concerning the analyzed fermentative interferents.

Phytochemical composition and in vitro safety evaluation of Ziziphora clinopodioides Lam. ethanolic extract: Cytotoxicity, genotoxicity and mutagenicity assessment

Ethnopharmacological relevanceThe application of the herb Ziziphora clinopodioides Lam. in folk medicine and as a food additive has been recommended due to its many claimed bioactivities. Regardless of the plant benefits, its safety considerations are largely unknown. Aim of the studyThe aim of the present research was to determine the chemical compositions and cytotoxicity, genotoxicity, and mutagenicity potentials of the ethanolic extract of Ziziphora clinopdioides Lam. (EEZC). Materials and methodsGC-MS and LC-MS analysis were used for chemical composition determination. The MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and trypan blue exclusion dye assays were used for cytotoxicity and the Comet assay was employed for genotoxicity assessment on human blood lymphocytes. Also, the Ames Salmonella/microsome test was carried out for the evaluation of mutagenicity. ResultsPulegone was the main component of the n-hexane fraction. Different phenolic acids and flavonoids were detected by LC-MS. The cytotoxicity study indicated a conspicuous decline in human lymphocyte viability ranging from 52% to 100% as showed by the MTT assay and 67% up to 100% by the trypan blue assay, at 1 and 10 mg/mL, respectively. The Comet assay results revealed a dose dependent genotoxicity, in so much as 90% and 98% of the cells were screened as damaged at concentrations of 5 and 10 mg/mL, respectively. An incidence rate of 8% and 13% of grade 4 damage was observed at 5 and 10 mg/mL, respectively. Additionally, the DNA damage index (DI) was elevated dose-dependently by a rising concentration of the extract, wherein the DI at 10 mg/mL concentration was 2.22, which was 22 times greater than that of negative control, and even more than positive control. The Ames test exhibited no signs of mutagenicity for neither Salmonella typhimurium TA98 nor TA100 strains, accompanied or unaccompanied by S9 metabolic activation. ConclusionResults indicated a dose-dependent cytotoxicity and genotoxicity potential of the EEZC on human lymphocytes, suggesting that this plant should be used with caution by consumers, even in the food and pharmaceutical industries. Since the plant usage in daily life continues to increase due to its ever growing phytotherapical and phytonutritional properties, it may pose a health risk by its high concentration's uptake. Although no mutagenicity of this extract was observed in this study, further research is recommended to clarify the mutagenic risks of this herb.