Relationships between cagA, vacA, and iceA genotypes of Helicobacter pylori and DNA damage in the gastric mucosa

Helicobacter pylori (H. pylori) is believed to dispose carriers to gastric cancer by inducing chronic inflammation. The inflammatory processes may result in the generation of reactive oxygen and nitrogen species that damage DNA. In this study, we investigated the relationships between DNA damage in the gastric mucosa and cogA, vocA, and iceA genotypes of H. pylori. The study was conducted with biopsies from the gastric antrum and corpus of 98 H. pylori-infected and 26 uninfected control patients. H. pylori genotypes were determined by PCR and DNA damage was measured in gastric mucosal cells by the Comet assay (single cell gel electrophoresis). All patients were nonsmokers, not abusing alcohol, and not using prescription or recreational drugs. Levels of DNA damage were significantly higher (P < 0.0001) in the H. pylori-infected patients than in uninfected patients. In comparison with the level of DNA damage in the uninfected controls, the extent of DNA damage in both the antrum (OR = 8.45; 95% Cl 2.33-37.72) and the corpus (OR 6.55; 95% Cl 2.52-17.72) was related to infection by cagA(+)/vocAs1m1 and iceA1 strains. The results indicate that the genotype of H. pylori is related to the amount of DNA damage in the gastric mucosa. These genotypes could serve as biomarkers for the risk of extensive DNA damage and possibly gastric cancer. (C) 2004 Wiley-Liss, Inc.

Oxidative stress status of highly prolific sows during gestation and lactation

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Desenvolvimento e validação de técnica quantitativa de análise de imagem para avaliação do teste do cometa corado pela prata

INTRODUÇÃO: O ensaio do cometa ou técnica da eletroforese de células isoladas é largamente empregado para avaliação de danos e reparo do DNA em células individuais. O material pode ser corado por técnicas de fluorescência ou por sal de prata. Este último apresenta vantagens técnicas, como o tipo de microscópio utilizado e a possibilidade de armazenamento das lâminas. A análise dos cometas pode ser feita de modo visual, porém há a desvantagem da subjetividade dos resultados, que pode ser minimizada por análise digital automatizada. OBJETIVOS: Desenvolvimento e validação de método de análise digital de cometas corados por sal de prata. MÉTODOS: Cinquenta cometas foram fotografados de maneira padronizada e impressos em papel. Além de medidas manualmente, essas imagens foram classificadas em cinco categorias por três avaliadores, antes e depois de pré-processadas automaticamente pelo software ImageJ 1.38x. As estimativas geradas pelos avaliadores foram comparadas quanto sua correlação e reprodutibilidade. em seguida, foram desenvolvidos algoritmos de análise digital das medidas, com base em filtros estatísticos de mediana e de mínimo. Os valores obtidos foram comparados com os estimados manual e visualmente após o pré-processamento. RESULTADOS: As medidas manuais das imagens pré-processadas apresentaram maior correlação intraclasse do que as imagens preliminares. Os parâmetros automatizados apresentaram alta correlação com as medidas manuais pré-processadas, sugerindo que este sistema aumenta a objetividade da análise, podendo ser utilizado na estimativa dos parâmetros dos cometas. CONCLUSÃO: A presente análise digital proposta para o teste do cometa corado pela prata mostrou-se factível e de melhor reprodutibilidade que a análise visual.

DNA damage in peripheral blood mononuclear cells of patients undergoing anti-tuberculosis treatment

Tuberculosis (TB), a chronic infectious disease, is a major cause of morbidity and mortality worldwide. Expression of iNOS and consequent production of NO during the inflammatory process is an important defense mechanism against TB bacteria. We have tested whether pulmonary TB patients undergoing anti-tuberculosis treatment present DNA damage, and whether this damage is related to oxidative stress, by evaluating total hydrophilic antioxidant capacity and iNOS expression. DNA damage in peripheral blood mononuclear cells from patients and healthy tuberculin test (PPD) positive controls was evaluated by single-cell gel electrophoresis (comet assay), and iNOS expression was measured by qPCR. We also evaluated total hydrophilic antioxidant capacity in plasma from patients and controls. Compared to controls, pulmonary TB patients under treatment presented increased DNA damage, which diminished during treatment. Also, the antioxidant capacity of these individuals was increased at the start of treatment, and reduced during treatment. TB patients showed lower iNOS expression, but expression tended to increase during treatment. Our results indicate that pulmonary TB patients under anti-TB treatment exhibit elevated DNA damage in peripheral blood mononuclear cells. This damage was not related to nitric oxide but may be due to other free radicals. (C) 2012 Elsevier B.V. All rights reserved.

DNA damage and nitric oxide production in mice following infection with L. chagasi

Leishmania chagasi, which causes visceral leishmaniasis in South America, is an obligate intracellular protozoan. Production of nitric oxide by macrophages during the inflammatory response is one of the main microbicidal mechanisms against this parasite. The goal of this study was to evaluate whether L. chagasi infection causes DNA damage in peripheral blood and spleen cells of Balb/c mice and whether such damage may be related to NO production. Balb/c mice were either infected with L chagasi or maintained as controls. The single-cell gel electrophoresis (comet) assay was used to measure DNA damage in peripheral blood and spleen cells, and the Griess reaction was used to measure NO production in the spleen. L chagasi infection induced DNA damage in peripheral blood and spleen cells of infected mice. Macrophages from the control group, challenged with L. chagasi, showed significantly (p < 0.05) greater NO production, compared to non-challenged cells. Treatment of spleen cells with N(G)-monomethyl-L-arginine (LNMMA) caused a significant reduction of NO production and DNA damage (p < 0.05). Our results indicate that L. chagasi induces DNA damage in the peripheral blood and spleen cells and that NO not only causes killing of the parasite but also induces DNA damage in adjacent cells. (C) 2011 Elsevier B.V. All rights reserved.

Biocompatibility in vitro tests of mineral trioxide aggregate and regular and white Portland cements

Mineral trioxide aggregate (MTA) and Portland cement are being used in dentistry as root end-filling materials. However, biocompatibility data concerning genotoxicity and cytotoxicity are needed for complete risk assessment of these compounds. In the present study, genotoxic and cytotoxic effects of MTA and Portland cements were evaluated in vitro using the alkaline single cell gel (comet) assay and trypan blue exclusion test, respectively, on mouse lymphoma cells. The results demonstrated that the single cell gel (comet) assay failed to detect DNA damage after a treatment of cells by MTA and Portland cements for concentrations up to 1000 mu g/ml. Similarly, results showed that none of the compounds tested were cytotoxic. Taken together, these results seem to indicate that MTA and Portland cements are not genotoxins and do not induce cellular death.

Evaluation of genetic damage in human peripheral lymphocytes exposed to mineral trioxide aggregate and Portland cements

Mineral trioxide aggregate (MTA) and Portland cement are being used in dentistry as root-end-filling material for periapical surgery and for the sealing of communications between the root canal system and the surrounding tissues. However, genotoxicity tests for complete risk assessment of these compounds have not been conducted up to now. In the present study, the genotoxic effects of MTA and Portland cements were evaluated in peripheral lymphocytes from 10 volunteers by the alkaline single cell gel (comet) assay. The results pointed out that the single cell gel (comet) assay failed to detect the presence of DNA damage after a treatment of peripheral lymphocytes by MTA and Portland cements for concentrations up to 1000 mu g mL(-1). In summary, our results indicate that exposure to MTA or Portland cements may not be a factor that increases the level of DNA lesions in human peripheral lymphocytes as detected by single cell gel (comet) assay.

Differential response related to genotoxicity between eggplant (Solanum melanogena) skin aqueous extract and its main purified anthocyanin (delphinidin) in vivo

Anthocyanins are the largest group of water-soluble pigments in the plant kingdom. A number of studies have demonstrated that anthocyanins present antioxidant capacity and show inhibitory effects on the growth of some cancer cells. Thus, the goal of this study was to evaluate both the antimutagenicity/antigenotoxicity and mutagenicity/genotoxicity of aqueous extract obtained from the Solanum melanogena, a possible novel source of anthocyanin, and its main purified anthocyanin extract (delphinidin), using the single cell (comet) assay and micronucleus test. Pretreatment with higher doses of the purified anthocyanin (10 and 20 mg/kg b.w.) led to a statistically significant reduction (p < 0.05) in the frequency of micronuclei in polychromatic erythrocytes induced by cyclophosphamide. The pattern of reduction ranged from 48% to 57% independent of concentration. No apparent: genotoxicity and mutagenicity was found for either the anthocyanin or delphinidin extracts. Taken together, these results suggest that mice pre-treated with specific compounds present in anthocyanins (delphinidin) displayed a lower incidence of mutations induced by cyclophosphamide. This finding emphasizes the potential of natural colorants to prevent mutations and also the applicability of genotoxic evaluation for improving health. Furthermore, the results presented here could be an additional argument to support the use of anthocyanins in the diet. (c) 2006 Published by Elsevier Ltd.

Genotoxicity of corrosion eluates obtained from endosseous implants

Purpose: Commercially pure titanium alloys are currently used as metallic biomaterials in implantology. Corrosion phenomena appear to play a decisive role in metallic implant long-term behavior. Thus, the goal of this study was to examine the genotoxic potential of corrosion eluates obtained from dental implants using Chinese ovary hamster cells in vitro by the single-cell gel (comet) assay. This technique detects deoxyribonucleic acid strand breaks in individual cells in alkaline conditions.Materials and Methods: the materials tested included 3 dental implants commercially available. Each of the tested materials was corroded in a solution consisting of equal amounts of acetic acid and sodium chloride (0.1 M) for 1, 3, 7, 14, and 21 days. The Chinese ovary hamster cultures were then exposed to all corrosion eluates obtained from endosseous dental implants for 30 minutes at 37 degrees C.Results: None of the eluates was found to exhibit genotoxicity, regardless of the type of dental implant used.Conclusion: the results suggest that all dental implants tested in this study did not induce deoxyribonucleic acid breakage as depicted by the single-cell gel (comet) assay.

Chlorhexidine induces DNA damage in rat peripheral leukocytes and oral mucosal cells

Objective: Chlorhexidine digluconate is widely used in dental practice for decreasing plaque control, controlling gingivitis and disinfecting root canals. However, the undesirable effects of chlorhexidine digluconate regarding its genotoxicity are conflicting in the literature. Thus, the aim of this study was to investigate the genotoxicity of chlorhexidine digluconate in rat peripheral blood and oral mucosal cells by the single cell gel (comet) assay and micronucleus assay.Methods: Thirty male Wistar rats were distributed into three groups: negative control; experimental group orally treated with 0.5 ml of 0.12% chlorhexidine digluconate, twice daily, during 8 days; and positive control, which received 4-nitroquinoline 1-oxide at 0.5 g/l by drinking water.Results: A statistically significant increase of DNA damage was observed in leukocytes and oral mucosal cells of the chlorhexidine digluconate treated group, as assessed by the comet assay. However, no increase of micronucleated cells was detected in reticulocytes from peripheral blood cells.Conclusions: Taken together, the data indicate that chlorhexidine digluconate is able to induce primary DNA damage in leukocytes and in oral mucosal cells, but no chromosome breakage or loss in erythrocytes.

Genotoxicity and cytotoxicity of glass ionomer cements on Chinese hamster ovary (CHO) cells

Glass ionomer cements are widely used in dentistry as restorative materials and adhesives for composite restorations. However, the results of genotoxicity studies using these materials are inconclusive in literature. The goal of this study was to examine the genotoxic and cytotoxic potential of three different glass ionomer cements available commercially (Ketac Cem, Ketac Molar and Vitrebond) by the single cell gel (comet) assay and trypan blue exclusion test, respectively. For this, such materials were exposed to Chinese hamster ovary (CHO) cells in vitro for 1 h at 37 degrees C. Data were assessed by Kruskall-Wallis nonparametric test. The results showed that the powder from Ketac Molar displayed genotoxicity only in the maximum concentration evaluated (100 mu g/mL). In the same way, the liquid from Vitrebond at 0.1% dilution caused an increase of DNA injury. Significant differences (P < 0.05) in cytotoxicity provoked by all powders tested of glass ionomer cements were observed for exposure at 1000 mu g/mL concentration. With respect to liquids of glass ionomer cements evaluated, the major toxic effect on cell viability was produced at 10%, beginning at the dilution of 0.5% for Vitrebond. Taken together, we conclude that some components of glass ionomer cements show both genotoxic and cytotoxic effects.

Lack of effect of prior treatment with fluoride on genotoxicity of two chemical agents in vitro

The goal of this study was to investigate the ability of fluoride to modulate the genotoxic effects induced by the oxidative agent hydrogen peroxide (H2O2) and the alkylating agent methyl methanesulfonate (MMS) in vitro by the single-cell gel ( comet) assay. Chinese hamster ovary cells were exposed in culture for 1 h at 37 degrees C to sodium fluoride at 7-100 mu g/ml. NaF-treated and control cells were then incubated with 0-10 mu M MMS in phosphate-buffered saline (PBS) for 15 min at 37 degrees C, or 7-100 mu M H2O2 in distilled water for 5 min on ice. Negative control cells were treated with PBS for 1 h at 37 degrees C. Clear concentration-related effects were observed for the two genotoxins. Increase of DNA damage induced by either MMS or H2O2 was not significantly altered by pretreatment with NaF. The data indicate that NaF does not modulate alkylation-induced genotoxicity or oxidative DNA damage as measured by the single-cell gel ( comet) assay. Copyright (c) 2007 S. Karger AG, Basel

Biocompatibility of glass-ionomer cements using mouse lymphoma cells in vitro

Glass-ionomer cements are widely used in dentistry as restorative materials and adhesives for composite restorations. A number of genotoxicity studies have been conducted using these materials with results conflicting so far. Thus, the approach was aimed to look at the genotoxic and cytotoxic potential of three different glass-ionomer cements available commercially (Ketac Cem, Ketac Molar and Vitrebond) by the single cell gel (comet) assay and trypan blue exclusion test, respectively. For this, such materials were exposed to mouse lymphoma cells in vitro for 1 h at 37 degrees C. Data were assessed by Kruskall-Wallis non-parametric test. The results showed that all powders assayed did not show genotoxic effects. on the other hand, the liquid from Vitrebond at 0.1% dilution caused an increase of DNA injury. Significant statistically differences (P < 0.05) in cytotoxicity provoked by all powders tested were observed for exposure at 1000 mu g mL(-1) concentration and 100 mu g mL(-1) for Ketac Molar. With respect to liquids of glass-ionomer cements evaluated, the major toxic effect on cell viability was produced at 1%, beginning at the dilution of 0.5% for Vitrebond. Taken together, these results support the notion that some components of glass-ionomer cements show both genotoxic and cytotoxic effects in higher concentrations.

No relationship between subchronic fluoride intake and DNA damage in Wistar rats

Fluoride has been widely used in dentistry because it is an effective caries prophylactic agent. However, excess fluoride may represent a hazard to human health, especially by causing injury on the genetic apparatus. Genotoxicity tests form an important part of cancer research and risk assessment of potential carcinogens. In the current study, the potential DNA damage associated with exposure to fluoride was assessed by the single cell gel ( comet) assay in peripheral blood, oral mucosa and brain cells in vivo. Male Wistar rats were exposed to sodium fluoride (NaF) at a 0, 7 and 100 ppm dose for drinking water during 6 weeks. The results pointed out that NaF did not contribute to the DNA damage in all cellular types evaluated as depicted by the mean tail moment and tail intensity. These findings are clinically important since they represent an important contribution to the correct evaluation of the potential health risk associated with dental agents exposure. Copyright (C) 2004 S. Karger AG, Basel.

Absence of DNA damage in multiple organs (blood, liver, kidney, thyroid gland and urinary bladder) after acute fluoride exposure in rats

Fluoride has been widely used in dentistry as a caries prophylactic agent. However, there has been some speculation that excess fluoride could cause an impact on genome integrity. In the current study, the potential DNA damage associated with exposure to fluoride was assessed in cells of blood, liver, kidney, thyroid gland and urinary bladder by the single cell gel (comet) assay. Male Wistar rats aging 75 days were distributed into seven groups: Groups 1 (control), 2, 3, 4, 5, 6 and 7 received 0 (deionized water), 10, 20, 40, 60, 80 and 100 mgF/Kg body weight from sodium fluoride (NaF), respectively, by gastrogavage. These groups were killed at 2 h after the administration of the fluoride doses. The level of DNA strand breaks did not increase in all organs evaluated and at all doses of NaF tested, as depicted by the mean tail moment. Taken together, our results suggest that oral exposure to NaF did not result in systemic genotoxic effect in multiple organs related to fluoride toxicity. Since DNA damage is an important step in events leading to carcinogenesis, this study represents a relevant contribution to the correct evaluation of the potential health risk associated with chemical exposure.