Deficiency in OGG1 protects against inflammation and mutagenic effects associated with H. pylori infection in mouse

BACKGROUND: Helicobacter pylori infection is associated with gastric cancer. Study with the Big Blue mouse model has reported a mutagenic effect associated with the H. pylori infection, as a result in part of oxidative DNA damage. The present work investigates the consequences of a deficiency in the OGG1 DNA glycosylase, responsible for the excision of 8-oxo guanine, on the inflammatory and genotoxic host response to the infection. MATERIALS AND METHODS: Big Blue Ogg1-/- C57BL/6 mice were orally inoculated with H. pylori strain SS1 or vehicle only, and sacrificed after 1, 3, or 6 months. The serologic response, histologic lesions, mutant frequency, and spectra of mutations were assessed in the stomach and compared to what observed in the wild-type (Wt) context. RESULTS: Inflammatory lesions induced in the gastric mucosa of H. pylori-infected mice, corresponding to a moderate gastritis, were less severe in Ogg1-/- than in Wt Big Blue mice. Analysis of antimicrobial humoral immunity exhibited a lower IgG2a serum level (Th1 response) after 6 months of infection in Ogg1-/- than in the Wt mice. In these conditions, the H. pylori-SS1 infection in the Ogg1-/- mice did not induce a mutagenic effect at the gastric epithelial cells level, either after 3 or 6 months. CONCLUSIONS: The inactivation of the OGG1 DNA glycosylase in mouse leads to less severe inflammatory lesions and abolished the mutagenic effect at the gastric epithelial cells level, induced by the H. pylori infection. These data suggest for the OGG1deficiency a protective role against inflammation and genotoxicity associated to the H. pylori infection.     

Toxicities of DDE on Wheat and Bioremediation of DDE by Fungus Pleurotus pulmonarius

An in vitro study was performed to determine the acute phytotoxicities and genotoxicity of DDE either spiked to soil or added to hydrophonic cultures on wheat Triticum aestivum. A 24-well plate was first used to determine toxicity on individual grains using conventional seed germination/seedling growth toxicity tests whereas a single cell electrophoresis system was applied to measure genotoxicity at single cell level for wheat. Hydrophonic cultures provide a simplified environment to screen for toxicities with high sensitivity. Inverse dose-response relationships were detected between exogenous DDE levels and one of the following parameters: seed germination, seedling growth, and genotoxicity. In contrast, soil reduced the stress on T. aestivum by lowering bioavailability leading to less DDE distributed in radicle and coleoptile, modulated growth, and enhanced tolerance. At all DDE doses spiked to soil including the reference safety level of 0.5 mg/kg, DNA breakage was detected in both radicle and coleoptile but their magnitudes did not correlate with the organ nor the soil DDE contents. Thus, although wheat is highly sensitive to the genotoxic effect of DDE, first demonstrated here, the seed germination test offers a simple quantitative measure of DDE's phytotoxicity in soil and hydrophonic cultures. This study also found that fungus Pleurotus pulmonarius, which secretes extracellular ligninolytic enzymes causing non-specific cleavage of lignin and organopollutants, remediated DDE spiked to soil. In 5 weeks, 78% of 10 mg/kg DDE was biodegraded, and the fungal-treated soil reduced acute toxicity on T. aestivum using the seed germination test.     

In silico assessment of chemical mutagenesis in comparison with results of Salmonella microsome assay on 909 chemicals

Genotoxicity is one of the important endpoints for risk assessment of environmental chemicals. Many short-term assays to evaluate genotoxicity have been developed and some of them are being used routinely. Although these assays can generally be completed within a short period, their throughput is not sufficient to assess the huge number of chemicals, which exist in our living environment without information on their safety. We have evaluated three commercially available in silico systems, i.e., DEREK, MultiCASE, and ADMEWorks, to assess chemical genotoxicity. We applied these systems to the 703 chemicals that had been evaluated by the Salmonella/microsome assay from CGX database published by Kirkland et al. [1]. We also applied these systems to the 206 existing chemicals in Japan that were recently evaluated using the Salmonella/microsome assay under GLP compliance (ECJ database). Sensitivity (the proportion of the positive in Salmonella/microsome assay correctly identified by the in silico system), specificity (the proportion of the negative in Salmonella/microsome assay correctly identified) and concordance (the proportion of correct identifications of the positive and the negative in Salmonella/microsome assay) were increased when we combined the three in silico systems to make a final decision in mutagenicity, and accordingly we concluded that in silico evaluation could be optimized by combining the evaluations from different systems. We also investigated whether there was any correlation between the Salmonella/microsome assay result and the molecular weight of the chemicals: high molecular weight (>3000) chemicals tended to give negative results. We propose a decision tree to assess chemical genotoxicity using a combination of the three in silico systems after pre-selection according to their molecular weight.     

Boric acid‐induced immunotoxicity and genotoxicity in model insect Galleria mellonella L. (Lepidoptera: Pyralidae)

Boric acid (BA) is widely used in various industrial process and can be accessed to nontarget organisms. This study aimed to investigate the insecticidal effects of BA and its toxic activities with respect to immunologic and genotoxic effects using Galleria mellonella larvae as a model. BA concentrations (78.125–10,000 ppm) were administrated to the larvae using the feeding method. Concentration‐dependent mortality was observed in all larval groups. Probit analysis revealed LC₃₀, LC₅₀, and LC₇₀ values to be 112.4, 320.1, and 911.4 ppm, respectively. These concentrations were used in all bioassays. Drastic reductions in total hemocyte counts along with changes in differential hemocyte counts were observed following BA treatment. Cell viability assays showed dose‐dependent reductions in viable cells and an increase in the necrotic and apoptotic ratios after BA treatment. However, mitotic indices of larval hemocytes did not change at all BA concentrations. The cytotoxic effect of BA led to a significant reduction in cellular immune responses such as encapsulation, melanization, and nodulation activities of treated larvae. While BA increased micronucleus ratios at the highest concentration, comet parameters indicating DNA damage increased in G. mellonella larval hemocytes at all concentrations. These report that BA suppresses the immune system of G. mellonella and also poses risks of genotoxicity at high concentrations.     

Low dose effects of chemicals as assessed by the flow cytometric in vivo micronucleus assay

Using flow cytometric automation of the mouse in vivo, micronucleus assay increases the sensitivity of the test. This is achieved through a very large increase in the number of cells scored, by a factor of 100×, which in turn greatly reduces the sampling error. With this method, dose–response relationships of in vivo micronucleus induction for four model agents mitomycin C (MMC), diepoxybutane (DEB), cyclophosphamide (CPA), and colchicine (COL) were studied at low dose levels. For the three clastogens MMC, DEB and CPA, linear dose–response relationships were found over the dose ranges studied, even in the very low dose region (defined as the dose region where the frequency of micronucleated erythrocytes is less than twice the baseline frequency). This is consistent with the view that no threshold should exist for genotoxic agents which target DNA. For COL a dose range was found, in which the frequency of micronucleated erythrocytes did not increase with dose, possibly indicating an in vivo threshold. The flow cytometric in vivo micronucleus assay represents one possibility for in vivo low dose–response studies.     

Assessment of the genotoxic potential of Hoechst 33342, SYBR-14 and PI using the SOS ChromoTest™

Abstract

Fluorochromes in combination with flow cytometry can be used for laboratory assessment of semen quality in humans and domestic animals. Some studies have reported the potential toxicity of these fluorochromes toward the cells analyzed, but not toward the laboratory technician who operates the analytical instrument. We tested the genotoxic potential of three fluorochromes, SYBR-14, propidium iodide, and Hoechst 33342, using the colorimetric SOS ChromoTest^?. The test revealed no genotoxic potential for any of the three fluorochromes within the dilution ranges investigated. We conclude that occasional direct contact with these fluorescent probes does not necessarily pose a genotoxic hazard.     

Genotoxicity assessment of Copaiba oil and its fractions in Swiss mice

Copaiba oil-resin, extracted from the trunk of Copaifera, and traditionally used in folk medicine in the treatment of various disorders, has been shown to be an effective antiinflamatory, antitumor, antitetanus, antiseptic and anti-blenorrhagea agent. As, there are few studies evaluating its genotoxicity, this aspect of the commercial oil-resin, and its volatile and resinous fractions, were evaluated in mice by comet assay and micronucleus (MN) test. A single dose of oil resin, volatile or resin fractions (500; 1,000 or 2,000 mg/kg b.w.) was administered by gavage. The chemical compositions of Copaiba oil resin and its fractions was analyzed by gas chromatography. According to comet assaying, treatment with either one did not increase DNA damage, and as to MN testing, there was no alteration in the incidence of micronucleated polychromatic erythrocytes. Chromatographic analysis of the oil-resin itself revealed sesquiterpenes, diterpenic carboxylic acid methyl esters and high levels of β-caryophyllene. Thus, it can be assumed that the oil resin and volatile and resinous fractions from the commercial product are not genotoxic or mutagenic.     

Toxicological Aspects of the Essential Oil from Cinnamodendron dinisii

The objective of this study was to determine cytotoxic activity, hemolytic activity, and to evaluate the ability of the essential oil from Cinnamodendron dinisii to induce DNA fragmentation of human lymphocytes. The essential oil was obtained by hydrodistillation. Cytotoxic activity was determined by the MTT method. Hemolytic activity was evaluated by spectrophotometric quantification of hemoglobin released by erythrocytes. Damage to lymphocyte DNA molecules was assessed by the Comet assay. The essential oil under study showed high cytotoxic activity on Vero cells (CC₅₀ = 35.72 μg/mL) and induced hemolysis in both hematocrits, besides leading to the oxidation of hemoglobin released. The genotoxic activity of C. dinisii essential oil was also observed, which induced concentration‐dependent DNA fragmentation of human lymphocytes and, at 50 μL/mL, it was more active than the positive control. The essential oil from C. dinisii has a toxic action, suggesting a special attention in the application of this oil to health‐promoting activities; however, among its components, there are molecules with potential for future application in anticancer therapies.     

Sodium Acetate,Sodium Acid Pyrophosphate,and Citric Acid Impacts on Isolated Peripheral Lymphocyte Viability,Proliferation, and DNA Damage

The present study examined the impacts of sodium acetate (SA), sodium acid pyrophosphate (SAPP), and citric acid (CA) on the viability, proliferation, and DNA damage of isolated lymphocytes in vitro. 3‐(4,5‐Dimethylthiazol‐2‐yl)‐2,5‐diphenyl tetrazolium bromide (MTT) and lactate dehydrogenase (LDH) release assays were adopted to evaluate cell viability, while comet assay was employed to assess the genotoxic effects. The cells were incubated with different levels of SA (50, 100, and 200 mM), SAPP (25, 50, and 100 mM/L), or CA (100, 200, and 300 μg/mL). The lymphocytes treated with the tested food additives showed concentration‐dependent decreases in both cell viability and proliferation. A concentration‐dependent increase in LDH release was also observed. The comet assay results indicated that SA, SAPP, and CA increased DNA damage percentage, tail DNA percentage, tail length, and tail moment in a concentration‐dependent manner. The current results showed that SA, SAPP, and CA are cytotoxic and genotoxic to isolated lymphocytes in vitro.