Application of comet assay to evaluate the impact of noise in DNA damage levels among exposed workers
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2022-04-01
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O ruído está associado a vários efeitos prejudiciais não auditivos cardiovasculares, e celulares, tais como a produção de espécies reativas de oxigénio que podem causar dano oxidativo no DNA. O presente projeto teve por objetivo a avaliação do dano oxidativo no DNA em sangue total, através do ensaio do cometa modificado com Fpg, em indivíduos ocupacionalmente expostos ao ruído. O estudo incluiu 56 trabalhadores de uma indústria metalúrgica, 44 dos quais envolvidos na produção (grupo exposto) e 12 em atividades administrativas ou semelhantes (grupo não-exposto). Os valores de exposição ao ruído foram obtidos pelo Departamento de Saúde Ambiental do Instituto Nacional de Saúde Doutor Ricardo Jorge, através de um procedimento interno acreditado, baseado na norma internacional ISO 9612:2011. Em simultâneo, foi avaliada a frequência cardiaca e de pressão vascular dos indivíduos. As condições a utilizar no ensaio do cometa, concretamente a concentração de Fpg, foram inicialmente otimizadas com um ensaio de titração em que foram testadas diferentes concentrações de Fpg (1.16, 1.33, 2.26 and 2.6 U/mL), para deteção do dano oxidativo de células mononucleares expostas a KBrO3 (0.5, 1.5, 4.5 mM). A metodologia otimizada foi utilizada para determinar os níveis de dano oxidativo no DNA em amostras de sangue total dos indivíduos incluídos no estudo. A análise dos resultados obtidos mostrou que não existiam diferenças estatisticamente significativas entre o grupo exposto e grupo não-exposto, nos níveis de dano oxidativo no DNA (p = 0.740), frequência cardíaca (p = 0.089), pressão arterial sistólica (p = 0.553), e pressão arterial diastólica (p = 0.271). Numa segunda abordagem, a amostra foi estratificada de acordo com os níveis de exposição ao ruído (≤85, entre 85 e 87, e ≥87 dB(A); valores de ação superior e limite de exposição, de acordo com a lei portuguesa); também neste caso, as diferenças entre os grupos no que diz respeito aos níveis de dano oxidativo no DNA, frequência cardíaca, pressão arterial sistólica, e pressão arterial diastólica não foram estatisticamente significativas (p = 0.794, p = 0.494, p = 0.995, e p = 0.499, respetivamente). Desta forma, o presente trabalho não provou haver uma relação entre a exposição ao ruído ocupacional e o aumento de dano oxidativo no DNA ou ocorrência de problemas cardiovasculares na amostra usada.
Noise is associated with many non-auditory harmful and cardiovascular effects such as vascular dysfunction, hypertension, leading to myocardial infarction, strokes, or heart failure and may cause oxidative DNA damage due to the release of ROS because of the stress. The present project aimed the evaluation of oxidative DNA damage in whole blood, through the Fpg-modified comet assay technique, in individuals exposed to occupational noise. This project included a sample of 56 workers of a metallurgic factory, 44 were working on the production (“Exposed” group), and 12 were performing administrative activities, or similar (“Non-exposed” group). The noise exposure values were obtained by the Environmental Health Department of the National Health Institute Doutor Ricardo Jorge under the accredited internal procedure based on the International Standard ISO 9612:2011. Simultaneously, was evaluated the heart rate and vascular pressure of the individuals. The comet assay conditions, specifically the Fpg concentration, were initially optimized with a titration assay where different Fpg concentrations (1.16, 1.33, 2.26 and 2.6 U/mL) where tested to allow the detection of oxidative damage on periferal blood mononuclear cells exposed to KBrO3 (0.5, 1.5, 4.5 mM). The optimized method was used to assess the oxidative DNA damage in the whole blood samples of the individuals included in the study. The analysis of the results showed that there were no statistically significant differences between the “Exposed” and the “Non-exposed” groups, regarding the oxidative DNA damage (p = 0.740), heart rate (p = 0.089), systolic blood pressure (p = 0.553), and diastolic blood pressure (p = 0.271). On a second approach, the sample was separated according to the exposure noise levels (≤85, between 85 and 87 e ≥87 dB(A); higher action and exposure limit values, acording to the portuguese law); also in this case, the differences between groups, regarding the oxidative DNA damage, heart rate, systolic and diastolic blood presures did not presented statistically significant differences (p = 0.794, p = 0.494, p = 0.995, e p = 0.499, respectively). Thus, the present work did not show correlation between occupational noise exposure and the increase of oxidative DNA damage or occurrence of cardiovascular issues in the present sample.
Noise is associated with many non-auditory harmful and cardiovascular effects such as vascular dysfunction, hypertension, leading to myocardial infarction, strokes, or heart failure and may cause oxidative DNA damage due to the release of ROS because of the stress. The present project aimed the evaluation of oxidative DNA damage in whole blood, through the Fpg-modified comet assay technique, in individuals exposed to occupational noise. This project included a sample of 56 workers of a metallurgic factory, 44 were working on the production (“Exposed” group), and 12 were performing administrative activities, or similar (“Non-exposed” group). The noise exposure values were obtained by the Environmental Health Department of the National Health Institute Doutor Ricardo Jorge under the accredited internal procedure based on the International Standard ISO 9612:2011. Simultaneously, was evaluated the heart rate and vascular pressure of the individuals. The comet assay conditions, specifically the Fpg concentration, were initially optimized with a titration assay where different Fpg concentrations (1.16, 1.33, 2.26 and 2.6 U/mL) where tested to allow the detection of oxidative damage on periferal blood mononuclear cells exposed to KBrO3 (0.5, 1.5, 4.5 mM). The optimized method was used to assess the oxidative DNA damage in the whole blood samples of the individuals included in the study. The analysis of the results showed that there were no statistically significant differences between the “Exposed” and the “Non-exposed” groups, regarding the oxidative DNA damage (p = 0.740), heart rate (p = 0.089), systolic blood pressure (p = 0.553), and diastolic blood pressure (p = 0.271). On a second approach, the sample was separated according to the exposure noise levels (≤85, between 85 and 87 e ≥87 dB(A); higher action and exposure limit values, acording to the portuguese law); also in this case, the differences between groups, regarding the oxidative DNA damage, heart rate, systolic and diastolic blood presures did not presented statistically significant differences (p = 0.794, p = 0.494, p = 0.995, e p = 0.499, respectively). Thus, the present work did not show correlation between occupational noise exposure and the increase of oxidative DNA damage or occurrence of cardiovascular issues in the present sample.
Descrição
Dissertação de Mestrado em Biotecnologia para as Ciências da Saúde
Palavras-chave
comet assay , Fpg