Made available in DSpace on 2016-01-26T18:55:42Z (GMT). No. of bitstreams: 1 Francijane Ferreira Paixao.pdf: 342552 bytes, checksum: 6c3ba89009e18cc2c819a6a07ee738d6 (MD5) Previous issue date: 2014-03-27 / This study aimed to evaluate the possible genotoxic effects of supplementation with capsiate extracted Pepper Capsicum annuum (sweet pepper), and the phenolic compound resveratrol extracted from grapes in Wistar rats by the micronucleus test in bone marrow. Sixty-four male rats, adult Wistar were divided into four groups (n = 16) and supplemented with: negative control group (GCN) - 0.5 ml of sodium chloride 0.9%; positive control group (GCP) - 50mg/kg cyclophosphamide; capsiate group (CG) - capsiate 10mg/kg diluted in 0.5 mL of 0.9% sodium chloride; resveratrol group (GR) - resveratrol 30mg/kg diluted in 0.5 mL of 0.9% sodium chloride. Only the PCM received intraperitoneal administration. The other groups were supplemented by gavage once a day for six weeks. At the end of six weeks the animals were anesthetized, sacrificed and the micronucleus test performed in bone marrow. The frequency of micronuclei in polychromatic erythrocytes of the BCM, PCM, GC and GR were 2.5, respectively, ± 0.53, 5.7 ± 1.03, 2.5 ± 0.73 and 2 ± 0.73 in a thousand cells. When comparing both groups there was statistically significant difference only GCP compared to other groups. It is concluded that supplementation at a dose of 10mg/kg capsiate and resveratrol at a dose of 30mg/Kg in daily frequency for 06 weeks did not produce genotoxicity in Wistar rats. / Este estudo teve como objetivo avaliar os possíveis efeitos genotóxicos da suplementação com o capsiate, extraído da pimenta Capsicum annuum (pimenta doce), e do composto fenólico resveratrol, extraído de uvas, em ratos Wistar através do teste de micronúcleo em medula óssea. Sessenta e quatro ratos machos, adultos de linhagem Wistar, foram distribuídos em quatro grupos (n=16) e suplementados com: grupo controle negativo (GCN)- 0,5ml de solução de cloreto de sódio 0,9%; grupo controle positivo (GCP)- 50mg/kg de ciclofosfamida; grupo capsiate (GC)- 10mg/kg de capsiate diluído em 0,5 mL de solução de cloreto de sódio 0,9%; grupo resveratrol (GR)- 30mg/kg de resveratrol diluído em 0,5 mL de solução de cloreto de sódio 0,9%. Somente o GCP recebeu a administração por via intraperitoneal. Os demais grupos foram suplementados por gavagem uma vez ao dia, durante seis semanas. Ao final das seis semanas os animais foram anestesiados, sacrificados e realizado o teste de micronúcleo em medula óssea. A frequência de micronúcleos em eritrócitos policromáticos do GCN, GCP, GC e GR foram, respectivamente, 2,5±0,53, 5,7±1,03, 2,5±0,73 e 2±0,73 em mil células. Na comparação entre os grupos observou-se diferença estatística significativa somente do GCP em relação aos demais grupos. Conclui-se que a suplementação de capsiate na dose de 10mg/Kg e resveratrol na dose de 30mg/Kg, em frequência diária, durante 06 semana, não produziu genotoxicidade em ratos Wistar.

Made available in DSpace on 2016-07-18T17:53:14Z (GMT). No. of bitstreams: 1 Francijane Ferreira Paixao.pdf: 342552 bytes, checksum: 6c3ba89009e18cc2c819a6a07ee738d6 (MD5) Previous issue date: 2014-03-27 / This study aimed to evaluate the possible genotoxic effects of supplementation with capsiate extracted Pepper Capsicum annuum (sweet pepper), and the phenolic compound resveratrol extracted from grapes in Wistar rats by the micronucleus test in bone marrow. Sixty-four male rats, adult Wistar were divided into four groups (n = 16) and supplemented with: negative control group (GCN) - 0.5 ml of sodium chloride 0.9%; positive control group (GCP) - 50mg/kg cyclophosphamide; capsiate group (CG) - capsiate 10mg/kg diluted in 0.5 mL of 0.9% sodium chloride; resveratrol group (GR) - resveratrol 30mg/kg diluted in 0.5 mL of 0.9% sodium chloride. Only the PCM received intraperitoneal administration. The other groups were supplemented by gavage once a day for six weeks. At the end of six weeks the animals were anesthetized, sacrificed and the micronucleus test performed in bone marrow. The frequency of micronuclei in polychromatic erythrocytes of the BCM, PCM, GC and GR were 2.5, respectively, ± 0.53, 5.7 ± 1.03, 2.5 ± 0.73 and 2 ± 0.73 in a thousand cells. When comparing both groups there was statistically significant difference only GCP compared to other groups. It is concluded that supplementation at a dose of 10mg/kg capsiate and resveratrol at a dose of 30mg/Kg in daily frequency for 06 weeks did not produce genotoxicity in Wistar rats. / Este estudo teve como objetivo avaliar os possíveis efeitos genotóxicos da suplementação com o capsiate, extraído da pimenta Capsicum annuum (pimenta doce), e do composto fenólico resveratrol, extraído de uvas, em ratos Wistar através do teste de micronúcleo em medula óssea. Sessenta e quatro ratos machos, adultos de linhagem Wistar, foram distribuídos em quatro grupos (n=16) e suplementados com: grupo controle negativo (GCN)- 0,5ml de solução de cloreto de sódio 0,9%; grupo controle positivo (GCP)- 50mg/kg de ciclofosfamida; grupo capsiate (GC)- 10mg/kg de capsiate diluído em 0,5 mL de solução de cloreto de sódio 0,9%; grupo resveratrol (GR)- 30mg/kg de resveratrol diluído em 0,5 mL de solução de cloreto de sódio 0,9%. Somente o GCP recebeu a administração por via intraperitoneal. Os demais grupos foram suplementados por gavagem uma vez ao dia, durante seis semanas. Ao final das seis semanas os animais foram anestesiados, sacrificados e realizado o teste de micronúcleo em medula óssea. A frequência de micronúcleos em eritrócitos policromáticos do GCN, GCP, GC e GR foram, respectivamente, 2,5±0,53, 5,7±1,03, 2,5±0,73 e 2±0,73 em mil células. Na comparação entre os grupos observou-se diferença estatística significativa somente do GCP em relação aos demais grupos. Conclui-se que a suplementação de capsiate na dose de 10mg/Kg e resveratrol na dose de 30mg/Kg, em frequência diária, durante 06 semana, não produziu genotoxicidade em ratos Wistar.

Premature cellular senescence is the process of permanent cell cycle arrest in response to various inducers, such as DNA damage, oxidative stress, chemotherapy agents, and irradiation. Senescent cells produce and secrete numbers of cytokines, chemokines, growth factors, which compose specific senescence-associated secretory phenotype (SASP). Senescence is considered to be an important barrier against tumor progression. On the other hand, senescent cells can also exert protumorigenic effects in their microenvironment. Based on this concept, the major aim of this thesis was to determine tumor cells senescence in terms of different inducers, namely chemotherapeutic agent docetaxel (DTX) and cytokines IFNγ and TNFα, and to demonstrate the role of immunotherapy in senescent cells elimination. Our results show that DTX-induced senescent cells can exert a tumor-promoting effect when co-injected with proliferating cells in mice. Importantly, we demonstrate that IL-12-based immunotherapy suppresses senescence-accelerated tumor growth. These results suggest that IL-12-based immunotherapy can be effectively used in anti-tumor therapy mainly in a case when the microenvironment is altered by the presence of tumor senescent cells. On the other hand, the data we obtained in vitro show that bystander or...

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In the tuberculosis patients, Mycobacterium tuberculosis can stimulate production of hydrogen peroxide in the host as a result of immune response. The H2O2 accumulate in pulmonary cells, causing oxidative stress that could lead to the cancer. We select TB patients for this study which investigates the effects of quercetin as there is an increased incidence of latent TB among the migrant population in the past few years and TB can increase the risk of cancer. Sperm and lymphocytes were treated with DNA damage inducers and quercetin (10µM, 25µM and 100µM), the responses evaluated using the Comet and micronucleus techniques. The gene expressions of COX1, COX2, P53 and Bcl-2 and catalase protein expression were investigated using the qPCR and Western blot techniques. The results showed that a substantial reduction of DNA damage in lymphocytes from TB patients and sperm from healthy donors from * P ≤ 0.0283 to *** P≤0.001in the Comet assay. In the MNi assay, the effect of quercetin in lymphocytes was more significant in reduce DNA damage, whereas the DNA damage induced by a food mutagen was significant, from *p 0.0405 to ***p 0.001. The qPCR showed significance down-regulation of COX1 and Bcl-2 gene expression, rated between *p 0.045 and **p 0.0074. However, the catalase protein was up-regulated by the nano form of quercetin when using lymphocytes from TB patients and showed significant changes at *p 0.0236. In conclusion, the nano form was found to be more efficient at the reduction of DNA damage in the Comet and micronucleus assays. Also, it down-regulated COX1 and Bcl-2 and up-regulated the catalase proteins indicating a possible role for quercetin, in genoprotection to TB through its enzyme modulating effect. / Libyan Embassy

No / Zinc oxide (ZnO) nanoparticles are widely used in cosmetics and sunscreens. Human epidermal keratinocytes may serve as the first portal of entry for these nanoparticles either directly through topically applied cosmetics or indirectly through any breaches in the skin integrity. Therefore, the objective of the present study was to assess the biological interactions of ZnO nanoparticles in primary human epidermal keratinocytes (HEK) as they are the most abundant cell type in the human epidermis. Cellular uptake of nanoparticles was investigated by scanning electron microscopy using back scattered electrons imaging as well as transmission electron microscopy. The electron microscopy revealed the internalization of ZnO nanoparticles in primary HEK after 6 h exposure at 14 microg/ml concentration. ZnO nanoparticles exhibited a time (6-24 h) as well as concentration (8-20 microg/ml) dependent inhibition of mitochondrial activity as evident by the MTT assay. A significant (p < 0.05) induction in DNA damage was observed in cells exposed to ZnO nanoparticles for 6 h at 8 and 14 microg/ml concentrations compared to control as evident in the Comet assay. This is the first study providing information on biological interactions of ZnO nanoparticles with primary human epidermal keratinocytes. Our findings demonstrate that ZnO nanoparticles are internalized by the human epidermal keratinocytes and elicit a cytotoxic and genotoxic response. Therefore, caution should be taken while using consumer products containing nanoparticles as any perturbation in the skin barrier could expose the underlying cells to nanoparticles.

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide. It has commonly been associated with exposure to tobacco-derived carcinogens and alcohol consumption. Pembrolizumab has shown to be effective in the treatment of many types of cancers such as melanoma, non-small cell lung cancer, due to its antiproliferative, immunoregulatory properties. The aim of this study was to investigate the effects of naked Pembrolizumab and Pembrolizumab liposome on the level of DNA damage, gene, and protein expressions in peripheral lymphocytes from HNC patients and compared to the healthy individuals by using the Comet and micronucleus assays. Western blotting and real-time polymerase chain reaction were performed to assess the potential of improving the repair mechanisms after treatment with naked Pembrolizumab and Pembrolizumab liposome. According to the results, Comet assay and micronucleus assay showed a significantly decreased DNA damage in the lymphocytes from HNC patients after being treated with naked Pembrolizumab and pembrolizumab liposome. Furthermore, the results have shown that naked Pembrolizumab and pembrolizumab liposomes (10 μg/ml) greatly decreased the oxidative stress produced by H2O2. Both forms of pembrolizumab have also demonstrated improving the repair mechanisms in lymphocytes from HNC patients by modulating the expression of P53, P21, and Bcl-2 at mRNA and protein levels. This study suggested that Pembrolizumab naked and liposome could have an antioxidant role alongside other actions in the treatment of HNSCC. However, further studies on Cancer cell lines and in vivo observation are required to validate the anticancer potential of pembrolizumab naked with liposome in HNC. / Ministry of Higher Education and Scientific Research, Libya

Submitted by Marcos Anselmo (marcos.anselmo@unipampa.edu.br) on 2017-06-12T14:17:48Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) JONATHALINE APOLLO DUARTE.pdf: 1033426 bytes, checksum: 65249cf69187d253ce4462e7152ce8db (MD5) / Approved for entry into archive by Marcos Anselmo (marcos.anselmo@unipampa.edu.br) on 2017-06-12T14:18:00Z (GMT) No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) JONATHALINE APOLLO DUARTE.pdf: 1033426 bytes, checksum: 65249cf69187d253ce4462e7152ce8db (MD5) / Made available in DSpace on 2017-06-12T14:18:00Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) JONATHALINE APOLLO DUARTE.pdf: 1033426 bytes, checksum: 65249cf69187d253ce4462e7152ce8db (MD5) Previous issue date: 2016 / O emprego de produtos naturais para o tratamento, cura ou prevenção de doenças pela população é datada desde os tempos mais remotos, e apesar dos inúmeros avanços na ciência, o uso de plantas com finalidade medicinal, ainda representa na maioria das vezes o único recurso terapêutico de muitas comunidades e de diversos grupos étnicos.Atualmente é possível verificarmos que apesar de todos os avanços nas indústrias farmacêuticas para a produção de fármacos sintéticos, a população ainda recorre às plantas medicinais.Entretanto, existe uma infinidade de plantas que ainda não são conhecidos os seus possíveis efeitos farmacológicos e/ou toxicológico. Diante desse contexto, a Schinus molle L. (Anacardiaceae), popularmente conhecida como anacauíta, é uma planta rica em óleo essencial, a qual vem sendo usada como uma opção de tratamento para diversas enfermidades, e apesar de seu amplo emprego na medicina popular, a literatura carece de informações voltadas para seu perfil imutoxicologico. Assim, investigouse os efeitos do óleo essencial frente a parâmetros citotóxico, mutagênico e genotoxico em cultura de linfócitos e macrófagos humanos. Inicialmente, determinaramse as DL50 para ambas as células estudadas, através do teste de proliferação celular, para então desenvolver os demais testes. As DL50 encontradas foram de 30.07μg/mL para linfócitos humanos e de 42.07μg/mL para macrófagos humanos, assim, foram definidas as concentrações a serem testadas, sendo essas DL50, DL50/10, DL50/100, DL50/1000 e DL50/10000 para as células em questão, e foi constatado que o óleo essencial foi capaz de promover citotoxicidade em concentrações superiores a DL50/1000, para ambas as células testadas. Entretanto, o mesmo proporciona efeito genotóxico em culturas de macrófagos, somente para as duas concentrações maiores e quando avaliado os frente a parâmetros mutagênicos, constatouse que, o mesmo não promove alterações cromossômicas assim como, também não alterou o índice de divisão celular, embora tenha sido capaz de proporcionar frequência de micronúcleo concentração dependente nos macrófagos. Contudo, é importante salientar a importância de conhecimento dos constituintes do óleo essencial da Schinus molle L., para maiores esclarecimentos referente a sua toxicidade, uma vez que, essa planta é amplamente empregada na medicina popular para as diversas finalidades. Dessa forma, os resultados encontrados nesse trabalho, tem a contribuir com a literatura. Para tanto, estudos complementares são necessários para auxiliar na construção completa do perfil toxicológico do óleo essencial da planta em estudo, buscando a segurança da população que a utiliza. / The use of natural products for the treatment, cure or prevention of disease by population is dated from the earliest times, and despite the numerous advances in science, the use of plants for medicinal purposes, yet is most often the only therapeutic resource many communities and various ethnic groups. It is currently possible to see that despite all the advances in the pharmaceutical industry for the production of synthetic drugs, people still make use of medicinal plants. However, there is a multitude of plants that are not yet known its possible pharmacological effects and / or toxicology. In this context, the Schinus molle L.(Anacardiaceae), popularly known as anacauíta, is a plant rich in essential oil, which has been used as a treatment option for various diseases, and despite its widespread use in therapy, the literature lacks information geared to your immunotoxicology profile. Thus, we investigated the effects of essential oil cytotoxic front parameters, mutagenic and genotoxic in cultured human lymphocytes and macrophages. Initially, the LD50 were determined for both the cells studied by the cell proliferation assay, and then to develop other tests. The LD50 was found to 30.07μg/ml for human lymphocytes and 42.07μg/ml for human macrophages, thus the concentrations to be tested have been identified and these LD50, LD50/10, LD50/100 LD50/1000 and LD50/10000 to the cells in question, and it was found that the essential oil was able to promote cytotoxicity at concentrations above LD50/1000, for both test cells, however, it provides genotoxic effects in macrophage cultures, only the two concentrations and larger when measured against the mutagens parameters, it was found that it does not promote chromosomal abnormalities as well as, did not alter the rate of cell division, although it was able to provide frequency micronucleus concentration dependent on macrophages. However, it is important to stress the importance of knowledge of essential oil constituents of Schinus molle L., for further information regarding its toxicity, since this plant is widely used in folk medicine for a variety of purposes. Thus, the results found in this work, is to contribute to the literature. Therefore, further studies are needed to help complete construction of the toxicological profile of the essential oil of the plant under study, seeking the safety of the population makes use of this. / Schinus molle L

Cyclotides are plant proteins with a unique topology, defined as the cyclic cystine knot motif. The motif endows cyclotides with exceptional chemical and biological stability. They also exhibit a wide range of biological activities including insecticidal, cytotoxic, anti-HIV and antimicrobial effects. Hence, cyclotides have become potential candidates in the development of peptide-based drugs; either as scaffolds to stabilize susceptible peptide sequences or as drugs by their own right. In this thesis, important parameters that could be inputs toward this development have been tuned. An extraction protocol that can be extended to industrial scale production of the cyclotides from natural sources was developed; accordingly, a single maceration with hydroalcoholic solutions of medium polarity represented an optimum extraction method. Moreover, it was shown that investigating the cyclotide content of cyclotide-bearing plants from diverse environments is a promising approach for extending the knowledge of both structural and biological diversity of these proteins. Five novel cyclotides with new sequence diversity were isolated and characterized from a violet that grows on Ethiopian highlands at an altitude of 3400 m. One of the areas where the cyclotide framework has attracted interest is the development of stable antimicrobial peptides. A stability study was carried out to determine the stability of the cyclotide framework in a cocktail of bacterial proteases and serum where the native forms of tested cyclotides exhibited high stability profile. Understanding the modes of cyclotide-cell interaction is certainly an important factor for the potential development of cyclotide-based drugs. Cellular studies were carried out using the comet assay and microautoradiography. A bell-shaped dose response curve was obtained for the DNA damaging effect of the cyclotides in the comet assay, which was the first toxicological assay of its kind on this class of proteins. The microautoradiography study revealed that the cyclotides penetrate into the cells even at cytotoxic concentrations. From previous reports, it was known that the cyclotides interact with membranes; the cellular studies in this thesis added to this knowledge by clearly demonstrating that these proteins have multiple modes of action.

The transcription factor NFAT5/TonEBP belongs to the Rel family, which also comprises NF ÛB and NFATc proteins. NFAT5 only shares structural and functional homology with other Rel family members at the level of the DNA binding domain, and differs from them considerably in other regions. NFAT5 enables mammalian cells to adapt to and withstand hypertonicity by orchestrating an osmoprotective gene expression program whose products include chaperones as well as ransporters and enzymes that increase the intracellular concentration of compatible osmolytes. NFAT5-null mice suffer severe embryonic and perinatal lethality, and surviving adults manifest growth defects, pronounced renal atrophy and lymphocyte dysfunction associated with ineffective responses to hypertonicity. To circumvent the lethality of these mice and study the function of NFAT5 in specific cell types without the possible side effects of generalized defects in the organism, we have produced conditional knockout mice that allow the deletion of NFAT5 in specific cell types. Here we have investigated the hypertonic stress response in wild-type and NFAT5-/- lymphocytes. Proliferating lymphocytes exposed to hypertonic conditions exhibited an early, NFAT5- independent, genotoxic stress-like response with induction of p53, p21 and GADD45, downregulation of cyclins E1, A2 and B1 mRNA, and arrest in S and G2/M. This was followed by an NFAT5-dependent adaptive phase in wild-type cells, which induced osmoprotective gene products, downregulated stress markers, and resumed cyclin expression and cell cycle progression. NFAT5-/- cells, however, failed to induce osmoprotective genes and though they downregulated genotoxic stress markers, they displayed defective cell cycle progression associated with reduced expression of cyclins E1, A2, B1, and aurora B kinase. Finally, T cell receptor-induced expression of cyclins, aurora B kinase, and cell cycle progression were inhibited in NFAT5-/- lymphocytes exposed to hypertonicity levels in the range reported in plasma in patients and animal models of osmoregulatory disorders. Our results support the conclusion that the activation of an osmoprotective gene expression program by NFAT5 enables cells to proliferate under hypertonic stress conditions by maintaining the expression of S and G2/M cyclins and cell cycle progression.