Made available in DSpace on 2015-01-26T13:21:21Z (GMT). No. of bitstreams: 0 Previous issue date: 2014-07-25Bitstream added on 2015-01-26T13:30:39Z : No. of bitstreams: 1 000795967.pdf: 970728 bytes, checksum: 1a9dc450a114117aa840a96d9f584e0e (MD5) / Nos últimos anos, tem crescido o interesse pela identificação de compostos naturais com potencial medicinal. O isotiocianato de alila (AITC), encontrado em plantas da família Cruciferae e composto majoritário da semente de mostarda, vem sendo avaliado quanto a sua atividade antineoplásica em bexiga devido à sua alta biodisponibilidade na urina a após ingestão. Neste estudo foram investigadas as atividades toxicogenética e toxicogenômica do AITC em linhagens celulares de carcinoma de bexiga com diferentes status do gene TP53 (RT4 – TP53 selvagem / T24 – TP53 mutado). As concentrações de AITC testadas nos ensaios de citotoxicidade, sobrevivência celular e clonogênica foram 0,005, 0,0625, 0,0725, 0,0825, 0,0925, 0,125 e 0,25 μM; nos testes do cometa, micronúcleo, ciclo celular, apoptose, morfologia e expressão gênica (ANLN, BAX, BCL-2, CDK-1, SMAD4, S100P e TP53) foram utilizadas as concentrações de 0,005, 0,0625, 0,0725, 0,0825 e 0,0925 μM. Os resultados mostraram aumento de danos primários no DNA (teste do cometa) para todas as concentrações testadas do AITC na linhagem T24 e para as concentrações mais altas (0,0725, 0,0825 e 0,0925 μM) na linhagem RT4; não foram observados aumentos nas frequências de células micronucleadas em ambas as linhagens. Nas análises de ciclo celular e apoptose foram observadas a parada do ciclo na fase G2/M para a linhagem T24 e altas taxas de apoptose precoce e necrose na linhagem RT4. Com relação à expressão gênica, houve aumento da expressão de S100P e BAX e diminuição da expressão de BCL-2 na linhagem RT4 e aumento da expressão de BCL-2, BAX e ANLN e diminuição da expressão de S100P na T24. Nenhuma alteração foi identificada para os genes SMAD4 e CDK1 nas duas linhagens tratadas com o AITC. Concluindo, o AITC foi capaz de induzir lesões primárias no DNA de células de ambas as linhagens de carcinoma de bexiga, mas que não se refletiram ... / Compounds obtained from fruits, vegetables and essential oils have been widely used to treat many diseases. The allyl isothiocyanate (AITC), also known as mustard essential oil, is found in plants of the cruciferous family and is abundant in the mustard seeds. Due to its high bioavailability in urine, AITC has been considered as a promising antineoplastic agent against bladder cancer. Therefore, the aims of this study were to investigate the toxicogenetic and toxicogenomic effects of AITC in the wild-type (RT4) and in a mutant (T24) TP53 bladder transitional carcinoma cell lines. AITC was tested at concentrations of 0.005, 0.0625, 0.0725, 0.0825, 0.0925, 0.125 and 0.25 μM in the cytotoxicity and cell and clonogenic survival assays; in the comet and micronucleus assays, flow cytometry, apoptosis and gene expression (ANLN, BAX, BCL-2, CDK-1, SMAD4, S100P e TP53) evaluations the concentrations of 0.005; 0.0625; 0.0725; 0.0825 e 0.0925 μM were used. The results showed increased primary DNA damage in T24 (0.005, 0.0625, 0.0725, 0.0825 and 0.0925 μM) and RT4 (0.0725, 0.0825 and 0.0925 μM) cells. However, no significant difference was detected in the frequency of micronucleated cells. A significant increase of cells at sub-G1 phase (0.0625, 0.0725 and 0.0825 μM) and significant decrease at S phase (0.005, 0.0625, 0.0725 and 0.0825 μM) were observed in RT4 cells after AITC treatments. In T24 cells, an increased sub-G1 rate (0.0725 and 0.0825 μM) and cell cycle arrest at G2 phase (0.0625, 0.0725 and 0.0825 μM) were detected. Furthermore, it was observed increased apoptosis and necrosis rates and increase of S100P and BAX and decrease of BCL-2 expression in the wild type TP53 cells. For the mutated TP53 cell line, increased expression of BCL-2, BAX and ANLN and decreased expression S100P were observed. No change was detected for the SMAD4 and CDK-1 genes. In conclusion, AITC was able to reach DNA and induce primary damage in both bladder ...

Orientador: Daysy Maria Fávero Salvadori / Coorientador: Glenda Nicioli da Silva / Banca: Luis Fernando Barbisan / Banca: Denise Crispim Tavares / Resumo: Nos últimos anos, tem crescido o interesse pela identificação de compostos naturais com potencial medicinal. O isotiocianato de alila (AITC), encontrado em plantas da família Cruciferae e composto majoritário da semente de mostarda, vem sendo avaliado quanto a sua atividade antineoplásica em bexiga devido à sua alta biodisponibilidade na urina a após ingestão. Neste estudo foram investigadas as atividades toxicogenética e toxicogenômica do AITC em linhagens celulares de carcinoma de bexiga com diferentes status do gene TP53 (RT4 - TP53 selvagem / T24 - TP53 mutado). As concentrações de AITC testadas nos ensaios de citotoxicidade, sobrevivência celular e clonogênica foram 0,005, 0,0625, 0,0725, 0,0825, 0,0925, 0,125 e 0,25 μM; nos testes do cometa, micronúcleo, ciclo celular, apoptose, morfologia e expressão gênica (ANLN, BAX, BCL-2, CDK-1, SMAD4, S100P e TP53) foram utilizadas as concentrações de 0,005, 0,0625, 0,0725, 0,0825 e 0,0925 μM. Os resultados mostraram aumento de danos primários no DNA (teste do cometa) para todas as concentrações testadas do AITC na linhagem T24 e para as concentrações mais altas (0,0725, 0,0825 e 0,0925 μM) na linhagem RT4; não foram observados aumentos nas frequências de células micronucleadas em ambas as linhagens. Nas análises de ciclo celular e apoptose foram observadas a parada do ciclo na fase G2/M para a linhagem T24 e altas taxas de apoptose precoce e necrose na linhagem RT4. Com relação à expressão gênica, houve aumento da expressão de S100P e BAX e diminuição da expressão de BCL-2 na linhagem RT4 e aumento da expressão de BCL-2, BAX e ANLN e diminuição da expressão de S100P na T24. Nenhuma alteração foi identificada para os genes SMAD4 e CDK1 nas duas linhagens tratadas com o AITC. Concluindo, o AITC foi capaz de induzir lesões primárias no DNA de células de ambas as linhagens de carcinoma de bexiga, mas que não se refletiram ... / Abstract: Compounds obtained from fruits, vegetables and essential oils have been widely used to treat many diseases. The allyl isothiocyanate (AITC), also known as mustard essential oil, is found in plants of the cruciferous family and is abundant in the mustard seeds. Due to its high bioavailability in urine, AITC has been considered as a promising antineoplastic agent against bladder cancer. Therefore, the aims of this study were to investigate the toxicogenetic and toxicogenomic effects of AITC in the wild-type (RT4) and in a mutant (T24) TP53 bladder transitional carcinoma cell lines. AITC was tested at concentrations of 0.005, 0.0625, 0.0725, 0.0825, 0.0925, 0.125 and 0.25 μM in the cytotoxicity and cell and clonogenic survival assays; in the comet and micronucleus assays, flow cytometry, apoptosis and gene expression (ANLN, BAX, BCL-2, CDK-1, SMAD4, S100P e TP53) evaluations the concentrations of 0.005; 0.0625; 0.0725; 0.0825 e 0.0925 μM were used. The results showed increased primary DNA damage in T24 (0.005, 0.0625, 0.0725, 0.0825 and 0.0925 μM) and RT4 (0.0725, 0.0825 and 0.0925 μM) cells. However, no significant difference was detected in the frequency of micronucleated cells. A significant increase of cells at sub-G1 phase (0.0625, 0.0725 and 0.0825 μM) and significant decrease at S phase (0.005, 0.0625, 0.0725 and 0.0825 μM) were observed in RT4 cells after AITC treatments. In T24 cells, an increased sub-G1 rate (0.0725 and 0.0825 μM) and cell cycle arrest at G2 phase (0.0625, 0.0725 and 0.0825 μM) were detected. Furthermore, it was observed increased apoptosis and necrosis rates and increase of S100P and BAX and decrease of BCL-2 expression in the wild type TP53 cells. For the mutated TP53 cell line, increased expression of BCL-2, BAX and ANLN and decreased expression S100P were observed. No change was detected for the SMAD4 and CDK-1 genes. In conclusion, AITC was able to reach DNA and induce primary damage in both bladder ... / Mestre