Yes / Exposure to DNA-damaging agents produces a range of stress-related responses. These change the expression of genes leading to mutations that cause cell cycle arrest, induction of apoptosis and cancer. We have examined the contribution of haploid and diploid DNA damage and genes involved in the regulation of the apoptotic process associated with exposure, The Comet assay was used to detect DNA damage and quantitative RT-PCR analysis (qPCR) to detect gene expression changes in lymphocytes and sperm in response to methyl methanesulfonate. In the Comet assay, cells were administered 0–1.2 mM of MMS at 37 °C for 30 min for lymphocytes and 32 °C for 60 min for sperm to obtain optimal survival for both cell types. In the Comet assay a significant increase in Olive tail moment (OTM) and % tail DNA indicated DNA damage at increasing concentrations compared to the control group. In the qPCR study, cells were treated for 4 h, and RNA was isolated at the end of the treatment. qPCR analysis of genes associated with DNA stress responses showed that TP53 and CDKN1A are upregulated, while BCL2 is downregulated compared with the control. Thus, MMS caused DNA damage in lymphocytes at increasing concentrations, but appeared not to have the same effect in sperm at the low concentrations. These results indicate that exposure to MMS increased DNA damage and triggered the apoptotic response by activating TP53, CDKN1A and BCL2. These findings of the processing of DNA damage in human lymphocytes and sperm should be taken into account when genotoxic alterations in both cell types are produced when monitoring human exposure. / Libyan Government

Metals and radionuclides are environmentally relevant contaminants, yet their potential impacts on marine organisms have not been adequately evaluated. This is especially true for exposures of longer duration and/or lower contaminant concentration (i.e. chronic) which are often more representative of real world scenarios. In this context, a suite of biomarkers at different levels of biological organisation were investigated in an ecologically relevant bivalve species, Mytilus galloprovin- cialis after exposure to nickel (a metal), zinc pyrithione (an organometal) and tritiated water (a radionuclide). These contaminants were chosen based on their differing properties, and hence, mechanisms of action. All three contaminants produced genotoxicity (DNA strand breaks, as measured by the comet assay, and induction of micronuclei [MN]). For nickel (> 1800 µg L −1 ) and tritiated water (15 MBq L−1 ), biomarkers at lower levels of biological organisation (i.e. DNA strand breaks, MN, changes in the expression of key stress response genes) were more sensitive than those at higher levels (i.e. clearance rate, attachment, tolerance of anoxia). In particular, exposure to tritiated water for 14 days resulted in DNA damage and molecular alterations without affecting higher level responses. As environmental contaminants could interact with other physical or chemical stressors in a complex environment, further exploration of biological responses revealed modulation by hyperthermia with concomitant changes in the transcriptional ex- pression of key defence genes (hsp70, hsp90, mt20, p53 and rad51). In contrast to nickel and tritiated water, exposure to both 0.2 and 2.0 µM zinc pyrithione caused significant deviation from concurrent controls for every biomarker examined, suggesting that further investigation of the environmental impacts of this contaminant is particularly necessary. Variation in biological responses induced by different contaminants suggests that potential links between levels of organisa- tion should be evaluated on a contaminant-specific basis. The integrated, multiple biomarker approach used in the current study provides a robust methodology for such studies, which could be translated to other ecologically relevant species for proper evaluation of risks to both environmental and human health.

Uranium has been mined for many years and used for fuel for nuclear reactors and materials for atomic weapons, ammunition, and armor. While the radioactivity associated with uranium mining has been linked to the development of lung and kidney cancers, and leukemia, little is known about the direct chemical genotoxicity of uranium. The overall hypothesis of the current research is that uranium can produce DNA damage by chemical genotoxicity mechanisms. Three specific aims were tested. In Aim 1, specific DNA lesions caused by direct interaction of uranium and DNA were investigated. Chinese Hamster Ovary cells (CHO) with mutations in various DNA repair pathways were exposed to 0 – 300 μM of soluble depleted uranium (DU) as uranyl acetate (UA) for 0 – 48 hr. Results indicate that UA readily enters CHO cells, with the highest concentration localizing in the nucleus. Clonogenics assay shows that UA is cytotoxic in each cell line with the greatest cytotoxicity in the base excision repair deficient EM9 cells and the nuclear excision repair deficient UV5 cells compared to the non-homologous end joining deficient V3.3 cells and the parental AA8 cells after 48 hr. This indicates that UA is forming DNA adducts that may be producing single strand breaks through hydrolysis rather than double strand breaks in CHO cells. Fast Micromethod® results indicate an increased amount of single strand breaks in the EM9 cells after 48 hr UA exposure compared to the V3.3 and AA8 cells. In Aim 2, the role of oxidative stress in producing DNA lesions was determined. Cellular oxidative stress has been implicated in the genotoxicity of many heavy metals as a mechanism of induced DNA damage. To investigate this possible mechanism, human bronchial epithelial cells (16HBE14o⁻) were exposed to 30 ppb (0.13 μM U) UA for 2 – 24 hr. UA did not significantly induce oxidative stress compared to untreated cells at 3 – 4 hr time points. These results suggest that cellular oxidative stress is not a major pathway of DU genotoxicity at low concentrations. In Aim 3, DNA damage response to uranium-induced DNA damage was investigated. It has been widely reported that metals can be genotoxic by inhibiting DNA repair. Cultured cells were co-exposed to 0.13 μM UA in the presence of 0 – 25 μM of etoposide for 0 – 48 hr. Results indicate that UA inhibited double strand break repair. Coexposures of etoposide and UA synergistically induced cytotoxicity compared to individual treatments and untreated cells. Co-exposed UA and etoposide treated 16HBE14o⁻ cells exhibited a decrease in phosphorylation of DNA repair proteins compared to etoposide treatments. Untreated and UA-treated 16HBE14o⁻ cells did not induce phosphorylation of DNA repair proteins. These results suggest that DU inhibits double strand break DNA repair at low concentrations in the presence of a known DNA double-strand damaging agent, etoposide. The inhibition of DNA repair by DU at environmentally relevant concentrations suggests a novel means by which uranium may exert its genotoxic effects. Results found at low dose exposures are not consistent with alterations seen with radioactivity, suggesting that the effects of uranium at low doses are due to its chemical genotoxic effects. Understanding how uranium reacts with DNA is important to better understand how this suspected carcinogen induces cancer and to help to elucidate mechanisms that produce cancers in people exposed to uranium.

The studies in this dissertation research were conducted to investigate the possible mode of action by which a brominated flame retardant, 2, 2-Bis (bromomethyl)-1, 3-propanediol (BMP) causes genotoxicity. Binding of BMP to DNA and BMP induced DNA strand breaks were investigated in SV-40 immortalized human uroepithelial cells (UROtsa) as an in vitro model for the bladder (a tissue that developed cancer after two year exposure to BMP in rodents). Results showed binding of [¹⁴C]-BMP equivalents to DNA increased with increased exposure time and concentration of [¹⁴C]-BMP. Comet analysis indicated BMP significantly increased the extent of DNA strand breaks at 1 and 3 h of incubation. However, strand breaks were repaired by 6 h of incubation. The DNA damaging effects of BMP at 1 h was concentration dependent. Compared with the parent compound, BMP-glucuronide (the predominant metabolite of BMP) bound less to DNA and produced less DNA strand breaks in UROtsa cells. Evidences that the BMP induced strand breaks were the result of an oxidative stress include: a concentration and time dependent increase in ROS generation; increased expression of Nrf2 and HSP70; complete attenuation of BMP induced DNA strand breaks by the antioxidant, NAC; and the presence of the oxidized base 8-OHguanine. UROtsa cells appear to be target cells for BMP because, as compared to rat hepatocytes (non-target cells), these cells lack the ability to detoxify BMP via glucuronidation and also because they are deficient in glutathione, a major intracellular antioxidant molecule. Both of these genotoxic events, DNA binding and oxidative DNA damage may, in part, contribute to BMP carcinogenicity observed in rodents. The relevance of current results to humans is remained to be established.

Amphibians are particularly sensitive to environmental degradation and, therefore, serve as effective environmental quality indicators. Research has suggested that amphibian declines are exacerbated by manmade environmental toxicants, especially those found in high concentrations in urban areas. The NIH has pinpointed genotoxicity as a major route of cancer causation, and has since developed stringent testing procedures for potentially hazardous chemicals. One such method, recognized for its simplicity and economy, is the micronucleus assay. A study was conducted assessing the genotoxicity of the widely used antimicrobial agent Triclosan to American Bullfrog tadpoles. Lithobates catesbeianus tadpoles were reared in glass aquaria containing ultra-high purity water and were dosed with nominal concentrations of 2.3 µg/L, 23 µg/L, and 230 µg/L Triclosan, reflecting 1x, 10x, and 100x concentrations of the compound as found in US surface waters. Eight replicates of each of the three levels of Triclosan contamination were prepared, as well as eight replicates per control group. Each replicate contained three tadpoles in a glass aquarium, from which one tadpole per tank was sampled after 1, 8, or 15 days following initial exposure to test compounds. Erythrocytes were prepared on slides and scored for micronucleus presence under 1000x magnification. Triclosan induced significant micronucleus formation after only 24 hours in all treatments relative to the negative control and exhibited a maximum of 15 micronuclei per 2,000 erythrocytes scored. Modeling of MN induction dynamics by treatment suggested that the best predictor of micronucleus induction was the acute TCS exposure level, as described by a linear mixed effects model including a binomial term of time exposed. Micronucleus induction was TCS concentration dose-dependent. This study supports that Triclosan induces significant genetic damage at environmentally relevant concentrations. It is clear that the effects of genotoxic agents must be certified so proper regulatory protocols can be developed and enforced, in order to conserve wildlife and promote human health.

O ensaio de frequência de micronúcleos é uma técnica importante utilizada para avaliar danos genotóxicos de agentes químicos ou físicos (como radiação ionizante) em células, baseado na quantificação de células contendo micronúcleos, que são fragmentos derivados de DNA danificado com coloração semelhante ao dos núcleos principais, mas com 5 a 30% do seu tamanho. Tradicionalmente, este ensaio é realizado usando microscopia de coloração convencional de acordo com Giemsa, mas atualmente esta técnica está sendo atualizada para uma abordagem automatizada que se baseia na dissolução da membrana plasmática por detergentes levando em conta apenas núcleos e micronúcleos com seu DNA corados por corantes fluorescentes, que pode discriminar núcleos de células viáveis dos das células inviáveis e permite a contagem por citometria de fluxo. Este trabalho avaliou a extensão de dano genotóxico radioinduzido por radiação gama (60Co) em doses entre 0,5 e 16Gy, e o potencial genotóxico de três peptídeos utilizados em medicina nuclear (PSMA-617, PSMA-11 e Ubiquicidina 29-41). As membranas celulares foram lisadas na presença dos corantes SYTOX® Green e EMA, e núcleos marcados com os dois fluorocromos foram considerados provenientes de células inviáveis e, portanto, removidos da análise. As quantidades de micronúcleos (porcentagem de eventos) nas amostras, foram proporcionais às doses de radiação, e puderam ser ajustadas a um modelo linear-quadrático (R2 = 0,993). Somente doses mais altas (8 e 16 Gy) e controles positivos induziram aumentos relevantes nas quantidades de micronúcleos. Os radiofármacos foram testados com concentrações de 0,1x, x, 10x e 100x a concentração utilizada em adultos, e não induziram dano em concentrações praticáveis. / The micronucleus frequency assay is an important technique used to evaluate genotoxic damage of chemical or physical agents (such as ionizing radiation) in cells, based on the quantification of micronucleus-containing cells, which are fragments derived from damaged DNA with similar coloration to of the main nuclei, but with 5 to 30% of its size. Traditionally, this assay is performed using conventional staining microscopy according to Giemsa, but currently this technique is being updated to an automated approach that relies on the dissolution of the plasma membrane by detergents taking into account only nuclei and micronuclei with their DNA stained by dyes fluorescence, which can discriminate nuclei from unviable or living cells and allows counting by flow cytometry. The genotoxic potential of three peptides used in nuclear medicine (PSMA-617, PSMA-11 and Ubiquicidine 29-41) was evaluated by gamma radiation (60Co) at doses in the range among 0.5 and 16Gy. Cell membranes were lysed in the presence of the SYTOX® Green and EMA dyes, and cores labeled with the two fluorochromes were considered to be from non-viable cells and therefore removed from the analysis. The amount of micronucleus (percentage of events) in the samples was proportional to the radiation doses, and could be adjusted to a linear-quadratic model (R2 = 0.993). Only higher doses (8 and 16 Gy) and positive control induced significant increases in micronucleus amounts. Radiopharmaceuticals were tested at concentrations of 0.1x, 1x, 10x, and 100x the concentration used in adults, and did not induce damage at feasible concentrations.

Orientador: Maria Aparecida Marin Morales / Banca: Silvia Tamie Matsumoto / Banca: Maria Tercília Vilela de Azeredo Oliveira / Resumo: O controle de plantas daninhas é fundamental para a produção agrícola, devido à sua competição pelos fatores limitantes de crescimento (nutrientes, água, luz) e aos seus efeitos alelopáticos. Pesticidas orgânicos sintéticos têm se tornado um importante elemento nas práticas de produção agrícola tecnificada, dando um grande impulso na indústria agroquímica em todo mundo. Os benefícios de tais produtos são claramente visíveis nos substanciais aumentos na produtividade. No entanto, o destino final desses compostos orgânicos, principalmente dos herbicidas, têm causado sérias preocupações, pelo grande volume utilizado. Preocupados principalmente com a possível interação desses produtos com o material genético dos organismos, nosso estudo visou compreender a ação da trifluralina, por ser este herbicida amplamente utilizado na agricultura. O presente estudo aborda avaliações toxicológicas, de mutagenicidade, genotoxicidade e carcinogenicidade do herbicida trifluralina. Também foram abordadas as características químicas, físicas, persistência, mobilidade, processo de degradação e mecanismos de ação do herbicida. Alguns dados foram obtidos junto às pesquisas realizadas na literatura científica disponível sobre o produto, mas a maioria das apresentações, aqui registradas, são decorrentes da própria investigação realizada com a aplicação do produto em organismos testes. Os dados obtidos foram comparados e discutidos, principalmente, com citações de autores especialistas no estudo da ação do produto e nas publicações da Agência de Proteção Ambiental Norte-Americana (USEPA). Aspectos da legislação brasileira dos agrotóxicos também foram sucintamente abordados / Abstract: The controlling of weed plants is fundamental for agricultural production, due to their competition by the limiting factors of growing (nutrients, water, light) and due to their alelopatics effects. Synthetic organic pesticides have becoming an important element in the practice of technical agricultural production, resulting in a great impulse in agrochemical industry around the world. The benefits from these products are clearly visible in the substantial increasing in the productivity. Otherwise, the final destination of these organic composts, mainly of the herbicides, have causing serious worrying because their high volume. Preoccupation mainly with the possible interaction of these products with the genetic organisms material, our study have the aim to examine the trifluralin action because this herbicide is widely utilized in the agriculture. The present work also looks at the toxicological interactions of mutagenicity, genotoxicity and carcinogenicity of the trifluralin herbicide as well the chemical and physical features, persistence, mobility, degradation process, and herbicide action mechanisms. Some data are from researches done in the available scientific literature about the product, but the majority of the presentation here reported is from our own investigation realized with the application of products in test organisms. The data collected were compared and discussed mainly with report of expert authors in the study of the action of the product and using publications in the United States Environmental Protection Agency (USEPA). Aspects of Brazilian agrotoxic legislation were also briefly discussed / Mestre

Orientador: Hélio Amante Miot / Banca: Paula Helena Ortiz Lima / Banca: Marco Antonio Rodrigues Fernandes / Resumo: O teste do cometa é utilizado para estimativa de dano ao DNA nos protocolos de avaliação do risco toxicológico. É rápido, de baixo custo, de fácil realização, seguro e pode ser aplicado em diversos tecidos. Corantes fluorescentes são os mais empregados para o ensaio. É também descrita a utilização de sais de prata com vantagens operacionais, carecendo de sistemas automatizados validados para esta análise. Neste trabalho, desenvolveram-se dois algoritmos automatizados para a análise do cometa corado pela prata. Foram realizados ensaios do cometa para sete grupos com dois camundongos Balb/c machos cada, submetidos ao tratamento intraperitoneal com soro fisiológico (G1), e três doses crescentes de genotóxicos conhecidos: MNU (N-nitroso-N-metilurea) (G2 - 5 μg/g, G3 - 25 μg/g, G4 - 50 μg/g) e DEN (N-nitrosodietilamina) (G5 - 20 μg/g, G6 - 80 μg/g, G7 - 180 μg/g). As lâminas dos testes do cometa foram confeccionadas a partir do sangue total e coradas pela prata e SYBR Gold. As coradas pela prata foram analisados por três avaliadores e pelos algoritmos automatizados. Aqueles corados pela fluorescência foram analisados pelo software Comet IV®. Os avaliadores apresentaram alta correlação de suas estimativas. Os algoritmos se correlacionaram fortemente com a análise dos avaliadores, principalmente com os escores obtidos pela classificação visual. Houve alta concordância na avaliação da sua repetitividade. Os algoritmos apresentaram resultados semelhantes, não indicando superioridade entre si para a análise do teste. Todos os métodos avaliados se mostraram capazes de diferenciar as concentrações dos genotóxicos utilizados. Porém, os algoritmos desenvolvidos não apresentaram correlação com os resultados obtidos pelo sistema do teste do cometa corado pela fluorescência, sugerindo que os métodos de coloração discriminem diferentes estruturas... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The comet assay is used to estimate DNA damage in the protocols for toxicological risk assessment. It's fast, inexpensive, easily performed, safe and can be applied in most tissues. Fluorescent dyes are frequently used for testing. Silver salts have been reported with some operational advantages; however automated systems for its analysis were not sufficiently validated for this staining. In this study, we have developed two algorithms for automated analysis of the comet assay silver stained. The assay was performed for seven groups of two (Balb/c) mice each, treated with intraperitoneal saline (G1) and three different genotoxic doses: MNU (N-nitroso-N-methylurea) (G2 - 5 μg/g, G3 - 25 μg/g, G4 - 50 μg/g) and DEN (N-nitrosodiethylamina) (G5 - 20 μg/g, G6 - 80 μg/g, G7 - 180 μg/g). The slides of the comet assay were made with total blood and were stained with silver and SYBR Gold. The silver stained slides were analyzed by three evaluators and the automated algorithms. Those stained by fluorescence were analyzed through software Comet IV®. Evaluators showed high correlation of their estimates. Both algorithms were correlated strongly with the evaluators' analysis mainly on the scores obtained by visual classification. Moreover, their agreement was high when assessed by repetitivity. The algorithms showed similar results, demonstrating no superiority to each other for the analysis of the test. All methods evaluated proved able of differentiating genotoxic concentrations used. However, the algorithms were not correlated with the results obtained by the automated system of comet assay fluorescence stained, suggesting that the staining methods linkage different genomic structures or with different affinities. The automated algorithms developed proved valid for the direct analysis of comet assay silver stained, allowing their use in genotoxicity research / Mestre

Submitted by Marcos Anselmo (marcos.anselmo@unipampa.edu.br) on 2016-09-22T14:32:37Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Gabriela de Souza da Silva.pdf: 747981 bytes, checksum: c098f507cfc2f2c6e90b43f8f1a81fec (MD5) / Approved for entry into archive by Marcos Anselmo (marcos.anselmo@unipampa.edu.br) on 2016-09-22T14:32:51Z (GMT) No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Gabriela de Souza da Silva.pdf: 747981 bytes, checksum: c098f507cfc2f2c6e90b43f8f1a81fec (MD5) / Made available in DSpace on 2016-09-22T14:32:51Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Gabriela de Souza da Silva.pdf: 747981 bytes, checksum: c098f507cfc2f2c6e90b43f8f1a81fec (MD5) Previous issue date: 2016-04-27 / O fluconazol é um fármaco antifúngico de amplo espectro muito utilizado pela população. Contudo, são poucos os registros na literatura que imputem a segurança do uso do fluconazol e tampouco a dose mínima capaz de induzir lesões no material genético do paciente. Embora a Agência Nacional de Vigilância Sanitária (ANVISA) exija testes de genotoxicidade para admissão de novos medicamentos, os que têm seu registro expedido antes do ano de 2006 não trazem essa obrigatoriedade, incluindo-se aqui o fluconazol. Portanto, é de extrema importância estudos sobre a avaliação genotoxicológica de fármacos, principalmente os que são tão conhecidos e utilizados como o fluconazol. O presente estudo investigou os efeitos genotóxicos da cápsula e do princípio ativo do fluconazol através do teste cometa e do teste de proliferação celular e também os efeitos citotóxicos da cápsula e do princípio ativo do fluconazol através do teste de viabilidade celular. As concentrações testadas da cápsula e do princípio ativo do fluconazol foram 6, 12, 30, 60 e 120 μg/mL. Todos os ensaios foram realizados em triplicatas e o peróxido foi utilizado como controle positivo e tampão PBS 7,4 como controle negativo. Todas as concentrações testadas da cápsula do Fluconazol (6, 12, 30, 60 e 120 μg/mL) demonstraram ser capazes de interferir negativamente no processo de proliferação celular, diminuindo o número de células proliferadas em todas as concentrações testadas, quando comparadas ao controle negativo. Já no caso do princípio ativo do Fluconazol, apenas as três últimas concentrações (30, 60 e 120 μg/mL) interferiram negativamente no processo de proliferação celular. Na avaliação do índice de dano ao DNA (teste cometa), foi observado dano à estrutura de DNA nas concentrações 60 e 120 μg/mL da cápsula do Fluconazol. Já as concentrações testadas com o princípio ativo do Fluconazol não foi observado dano à estrutura de DNA. Foi verificado que a cápsula do Fluconazol possui atividade citotóxica sobre leucócitos humanos nas duas maiores concentrações testadas (60 e 120 μg/mL) de forma concentração dependente. Porém, o princípio ativo do Fluconazol não demonstrou ser citotóxico nas concentrações testadas (6 12, 30, 60 e 120 μg/mL). / Fluconazole is a broad-spectrum antifungal drug widely used by the population. However, there are few reports in the literature that imput the safety of the use of fluconazole and either the minimum dose capable of inducing lesions in the genetic material of the patient. Although the National Health Surveillance Agency (ANVISA) requires genotoxicity tests for admission of new medicines, which have their registration issued before 2006 do not bring this obligation, including here fluconazole. Therefore, it is extremely important genotoxicológica studies on the evaluation of drugs, especially such that are known and used as fluconazole. This study investigated the genotoxic effects of the capsule and the active principle of fluconazole by the comet test and the cell proliferation test and also the cytotoxic effects of the capsule and the active principle of fluconazole by cell viability test. The tested concentrations of the capsule and the active principle of fluconazole were 6, 12, 30, 60, and 120 mg/mL. All assays were performed in triplicate and peroxide was used as a positive control and PBS buffer as negative control 7.4. All tested concentrations of fluconazole capsule (6, 12, 30, 60 and 120 μg/mL) were capable of induce a negative interference in the cellular proliferation process, reducing the number of proliferating cells at all concentrations tested when compared to the negative control. In the case of the active ingredient, the fluconazole, only the last three concentrations (30, 60, and 120 μg/mL) were able to decrease the cellular proliferation process. In the assessment of DNA damage index (comet assay), there was observed damage of fluconazole capsule on the DNA structure at concentrations 60 and 120 μg/mL. On the other hand, the concentrations tested with the active ingredient of Fluconazole was not observed damage to the DNA structure. Fluconazole has been found that the capsule has a cytotoxic activity on human leukocytes at the two highest concentrations tested (60 and 120 μg/mL) as a concentration-dependent manner. However, the active ingredient of Fluconazole not induced any cytotoxic effect at the concentrations tested (6, 12, 30, 60, and 120 μg/mL).

Submitted by Marcos Anselmo (marcos.anselmo@unipampa.edu.br) on 2016-09-22T18:11:33Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) MARIA FERNANDA DE MOURA LEÃO.pdf: 2846166 bytes, checksum: f1e8c6a8d84f559be79cb61300900f00 (MD5) / Approved for entry into archive by Marcos Anselmo (marcos.anselmo@unipampa.edu.br) on 2016-09-22T18:12:09Z (GMT) No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) MARIA FERNANDA DE MOURA LEÃO.pdf: 2846166 bytes, checksum: f1e8c6a8d84f559be79cb61300900f00 (MD5) / Made available in DSpace on 2016-09-22T18:12:09Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) MARIA FERNANDA DE MOURA LEÃO.pdf: 2846166 bytes, checksum: f1e8c6a8d84f559be79cb61300900f00 (MD5) Previous issue date: 2016 / A hipertensão arterial é uma condição clínica de ocorrência multifatorial, sendo a mais frequente entre as doenças crônicas não transmissíveis no Brasil. É caracterizada pelo aumento e manutenção dos níveis pressóricos acima de 140mmHg (pressão sistólica) e 90mmHg (pressão diastólica). É também o maior agravante nas complicações cardiovasculares e está associada ao surgimento de outras co-morbidades. A OMS estima que 40% da população mundial acima dos 25 anos seja hipertensa, estando localizada majoritariamente em países onde a renda e o nível de escolaridade são menores. Depois de diagnosticada, a hipertensão arterial deve ser tratada com mudanças no estilo de vida e medicamentos que mantenham os níveis pressóricos controlados, sendo assim, várias classes de anti-hipertensivos são usados nesse tratamento. Visando ampliar o acesso a medicamentos básicos e permitir uma melhor adesão ao tratamento, o Governo Federal criou em 2004 o Programa Farmácia Popular do Brasil, uma parceria entre governo e instituições públicas e privadas para fornecer a população medicamentos para o controle da hipertensão, diabetes, asma, dislipidemias, anticoncepcionais, entre outros; de forma gratuita ou subsidiada em até 90% do valor. Para tratar a hipertensão, o Programa Farmácia Popular do Brasil distribui de forma gratuita os anti-hipertensivos – atenolol, captopril, cloridrato de propranolol, hidroclorotiazida, losartana e maleato de enalapril, todos estes sendo lançados no mercado anterior ao ano de 2004, quando testes de segurança genética ainda não eram obrigatórios, de acordo com a Resolução nº 90/2004 da ANVISA. Desta forma, o objetivo deste trabalho foi avaliar o potencial genotóxico de anti-hipertensivos distribuídos pela Farmácia Popular em células leucocitárias humanas, haja vista que são fármacos amplamente utilizados e se faz necessários maiores estudos que garantam a segurança genotoxicológica dos mesmos. Os testes foram desenvolvidos a partir de culturas celulares tratadas com cinco diferentes concentrações dos anti-hipertensivos, sendo avaliados parâmetros genotoxicológicos – viabilidade celular e índice de dano ao DNA (teste Cometa), e parâmetros mutagênicos – teste de micronúcleo e instabilidade cromossômica numérica. Os resultados mostram que captopril e maleato de enalapril foram capazes de diminuir a viabilidade celular e causar danos ao DNA conforme o aumento da dose, e que, o atenolol foi capaz de também diminuir a viabilidade celular de acordo com o aumento da dose. A hidroclorotiazida também mostrou causar dano ao DNA, porém esse dano ocorreu de forma igual para as cinco doses. Quanto aos parâmetros mutagênicos, nenhum dos seis anti-hipertensivos testados e, em nenhuma das cinco concentrações foi capaz de causar alterações mutagências. / Hypertension is an clinical condition of the multifactorial occurrence, the most frequent among chronic diseases not transferable in Brazil. It is characterized by increase and maintenance blood pressure levels above 140mmHg (sistolic pressure) and 90mmHg (diastolic pressure). It i also the most aggravating in the cardiovascular complications and it is associated with the appearance of others comorbidities. The WHO estimes that 40% of the world population over 25 years old is hypertensive, being located mostly in countries where income and educations levels are lower. After the diagnosed, hypertension must be treated with changes in the life style and drugs that maintain pressure levels controled, therefore, several classes of antihypertensive are used in this treatment. Aiming to expand access to basic drugs and allow a better adhesion to treatment, the federal government created in 2004 the “Farmácia Popular do Brasil” Program, an association among government and public and private institutions to provide the people medications for control of the hypertension, diabetes, asthma, dyslipidemia, contraceptives, etc, free or subsidized form by up to 90% of the value. To treat hypertension, the “Farmácia Popular do Brasil” Program distributes of free form antihypertensive – atenolol, captopril, propranolol hydrochloride, hydrochlorotiazhide, losartan and enalapril maleate, all these being released before the year 2004, when genetic safety tests still not required, according to Resolution nº 90/2004 of ANVISA. Thus, the objective this work was to evaluate the genotoxic potencial of the antihypertensives distributed by “Farmácia Popular” in human leukocytes cells, considering that they are widely used drugs and it is necessary larger studies that ensure the genotoxicology their safety. The tests are desenvolved from cell cultures treated with five different concentrations of the antihypertensives, being evaluated genotoxicological parameters – cell viability and DNA damage index, and mutagenic parameters – micronucleus test and numerical chromossomal aberrations. The results showed that captopril and enalapril maleate were able to reduce cell viability and cause damage to DNA wiht increasing dose. The hydrochlorothiazide also showed to cause DNA damage, but, this damage occorred equallity for five doses. As for mutagenic parameters , none of the six antihypertensives tested and, none of the five concentrations were capable to cause mutagenic changes.