A carcinogênese é um processo longo que envolve múltiplas etapas na transformação das células normais em malignas. Duas hipóteses têm sido propostas para explicar o potencial heterogêneo de células neoplásicas e o processo de desenvolvimento de neoplasias: o modelo estocástico, em que uma população distinta de células neoplásicas adquire um conjunto de mutações somáticas e desenvolve capacidade metastática e o modelo hierárquico, no qual neoplasias primárias e suas metástases são iniciadas por um número pequeno de células, conhecidas \"cancer stem cells\" (CSC). Assim, o HCC poder ser originado de células tronco/progenitoras hepáticas ou células ovais e hepatócitos. Aventou-se a hipótese neste trabalho que a BI reduz o número de células ovais hepáticas, regulando proteínas (CK19, β-catenina e ALDH1A1) que possam estar relacionadas ao papel destas células na hepatocarcinogênese. Ratos Wistar foram submetidos ao modelo de hepatocarcinogênese do \"Hepatócito Resistente\" e receberam durante 4 semanas consecutivas: 16mg/100g p.c. de β-ionona (BI) ou somente 0,25mL/100g p.c. de óleo de milho (grupo controle - OM). Estes animais foram distribuídos em 4 pontos de eutanásia (P0, P1, P2 e P3 - 7, 14, 21 e 35 dias após a HP respectivamente), para avaliação da cinética das células ovais hepáticas. Foi observado que o tratamento com BI diminuiu a presença de células ovais GST-P positivas e reduziu o percentual de LPN positivas para ALDH1A1, β-catenina e CK19 quando comparado ao do respectivo grupo controle OM. Em conclusão, o tratamento com BI reduziu o número das células ovais, bem como os níveis de proteínas a elas relacionadas e que conferem ao HCC mau prognóstico e maior agressividade. / Carcinogenesis is a long process that involves multiple phases in the transformation of normal cells into malignant. Two hypotheses have been proposed to explain the heterogeneous potential of neoplastic cells and the process of carcinogenesis: a stochastic model in which a distinct population of neoplastic cells acquires a set of somatic mutations and develops metastatic capacity and hierarchical model in which the primary tumors and their metastases are initiated by a small number of cells, known \"cancer stem cells\" (CSC). Therefore, the HCC can be originate from stem/progenitor cells or hepatic oval cells and hepatocytes. Ventured the hypothesis in this work that β-ionone (BI) reduces the number of hepatic oval cells, regulating proteins (CK19, β-catenin and ALDH1A1) that may be related to the role of these cells in hepatocarcinogenesis. Wistar rats were submitted to hepatocarcinogenesis model of \"Resistant Hepatocyte\" and received for 4 consecutive weeks: 16mg/100g body weight of BI or 0,25 mL/100g body weight of corn oil (control group - CO). These animals were euthanized at 4 points (P0, P1, P2 and P3 - 7, 14, 21 and 35 days after PH, respectively) to evaluate the kinetics of hepatic oval cells. It was observed that the treatment of BI decreased the presence of GST-P oval cells positive and reduced the percentage of positive ALDH1A1 LPN, β-catenin and CK19 compared to the respective control group OM. In conclusion, the treatment with BI reduced the number of oval cells as well as the levels of proteins related to them and gives a poor prognosis in HCC and increased aggressiveness.

No presente estudo avaliou-se a atividade quimiopreventiva da tributirina (T), pró-fármaco do ácido butírico (AB) e presente naturalmente em leite e derivados, quando administrada a ratos Wistar durante as etapas de iniciação e seleção/promoção do modelo de hepatocarcinogênese do \"Hepatócito Resistente\" (RH). Os animais receberam diariamente, durante 8 semanas consecutivas e por entubação gástrica: T (200 mg/100 g de peso corpóreo [p.c.]; grupo TB) ou maltodextrina (300 mg/100 g de p.c.; grupo MD; controle isocalórico). Duas semanas após o início dos tratamentos, os grupos foram submetidos ao modelo do RH. Esse consistiu na aplicação intraperitoneal de uma dose do agente iniciante dietilnitrosamina (DEN, 20 mg/100 g de p.c.), seguida, 2 semanas após, da aplicação de 4 doses consecutivas de 2-acetilaminofluoreno (AAF; 2 mg/100 g de p.c.) e de uma hepatectomia parcial (HP) a 70%, acrescida de 2 doses de AAF (2 mg/100 g de p.c.) 2 e 4 dias após a cirurgia. Decorridas 6 semanas após a iniciação com DEN, os animais foram eutanasiados administrando-se, entretanto, 2 horas antes desse procedimento 5-bromo-2-desoxiuridina (BrdU) (10 mg/100 g de p.c.). De acordo com a análise macroscópica dos fígados, e em comparação ao grupo MD, verificou-se que o grupo TB apresentou menor multiplicidade (p &lt; 0,05) e tamanho (p &lt; 0,05) de lesões pré-neoplásicas (LPN) hepáticas. Em relação à análise morfométrica das LPN hepáticas positivas para a enzima glutationa S-transferase forma placentária (GST-P) totais (persistentes + em remodelação) e também nas LPN hepáticas GST-P positivas persistentes, observou-se que em comparação ao grupo MD, o TB apresentou menor tamanho (p &lt; 0,05) e área do corte ocupada (p &lt; 0,05) por essas lesões. O grupo TB apresentou maior porcentagem (p &lt; 0,05) de LPN hepáticas GST-P positivas em remodelação e maior número (p &lt; 0,05) de corpúsculos apoptóticos (CA) nessas LPN em comparação ao MD. Não houve diferenças (p &gt; 0,05) entre os grupos MD e TB em relação à proliferação celular e aos danos no DNA. De acordo com a imunoistoquímica para p53, o grupo TB apresentou menor porcentagem (p < 0,05) de LPN hepáticas totais e persistentes positivas para a proteína, comparado ao MD. O grupo TB apresentou menor ativação (p &lt; 0,05) do fator nuclear-kB (NF-kB) quando comparado ao MD. O grupo TB tendeu a apresentar maior acetilação de histona H3 resíduo de lisina 9 (H3K9) em relação ao grupo MD. A T apresentou atividade quimiopreventiva promissora quando administrada a ratos Wistar diariamente e durante 8 semanas, abrangendo as etapas de iniciação e seleção/promoção do modelo de hepatocarcinogênese do RH. Inibição de LPN hepáticas GST-P positivas persistentes, indução da remodelação e da apoptose, além de normalização da função de p53, inibição da ativação do NF-kB e indução da acetilação de H3K9 parecem estar envolvidas com as ações anticarcinogênicas da T. / Chemopreventive activity of tributyrin (T), a butyric acid prodrug found in milk and its derivatives, was evaluated during the initial phases of the \"Resistant Hepatocyte\" (RH) model of hepatocarcinogenesis. During 8 consecutive weeks, rats received T (200 mg/100 g body weight; TB group) or maltodextrin (M; 300 mg/100 g body weight; MD group; isocaloric control). Two weeks after the beginning of the treatments, the animals received one dose of diethylnitrosamine (DEN; 20 mg/100 g body weight) for initiation. Two weeks later, the animals received six doses of 2-acetylaminofluorene (AAF, 2 mg/100 g body weight), 4 consecutive doses before partial (2/3) hepatectomy and the remaining, two and four days after surgery. All animals were euthanized 6 weeks after DEN administration. Two hours before euthanasia, rats received 5-bromo-2-desoxiuridina (BrdU) (10 mg/100 g body weight). Multiplicity and area of visible hepatocyte nodules/animal were smaller (p &lt; 0,05) in TB group, compared to MD group. Mean area and % liver section area occupied by total (persistant + remodeling) hepatic placental glutathione S-transferase (GST-P) positive preneoplastic lesions (PNL) and persistant hepatic GST-P positive PNL were smaller (p <&lt; 0,05) in TB group, compared to control group. Compared to MD group, TB group presented increased (p &lt; 0,05) % of remodeling hepatic GST-P positive PNL and increased number of apoptotic bodies in these PNL. No differences (p gt; 0,05) were observed between MD and TB groups regarding cell proliferation and DNA damage. TB group presented reduced (p &lt; 0,05) % of total and persistant PNL p53 positive compared to MD group. TB group presented reduced (p < 0,05) nuclear factor-kB (NF-kB) activation in comparison with MD group. Lysine 9 acetylation site in H3 (H3K9) tended to be increased in TB group compared to MD. These results indicate that T represents promising chemopreventive agent against hepatocarcinogenesis when administered to Wistar rats during 8 consecutive weeks on initial phases of RH model. Persistant hepatic GST-P positive PNL inhibition, remodeling and apoptosis induction, p53 function normalization, inhibition of NF-kB activation and H3K9 acetylation can be involved with antineoplastic T actions.

O carcinoma hepatocelular (HCC) apresenta mau prognóstico o que torna importante sua quimioprevenção. Nesse sentido, a tributirina (TB), um inibidor de desacetilases de histonas (HDACi), mostrou-se um quimiopreventivo promissor da hepatocarcinogênese. Avaliaram-se aqui efeitos quimiopreventivos de lipídios estruturados (EST) obtidos por interesterificação enzimática a partir da TB com o óleo de linhaça (LIN). Ratos foram tratados com EST (grupo EST; 165 mg/100g peso corpóreo [p.c]), TB (grupo TB; 200 mg/100g p.c), LIN (grupo LIN; 133 mg/100g p.c), mistura de TB com LIN (grupo LIN; 165 mg/100g p.c) ou maltodextrina (MD) (grupo MD; controle isocalórico; 300 mg/100g p.c) diariamente durante 8 semanas consecutivas por gavagem. Duas semanas após início dos tratamentos, os animais foram submetidos ao modelo de hepatocarcinogênese do \"hepatócito resistente\" (RH). Os grupos EST e TB apresentaram atividade quimiopreventiva bloqueadora e supressora, respectivamente, da hepatocarcinogênese. TB induziu a apoptose, ao contrário dos EST. O tratamento com TB resultou na acetilação e trimetilação da H3K9 e H3K27, enquanto EST atuaram somente na trimetilação das mesmas. Quando analisada a expressão de genes envolvidos com modificações em histonas, EST e TB reduziram a expressão de Ezh2 e de Hdac4. Por outro lado, somente os EST aumentaram a expressão de Hdac6. Tal efeito por parte dos EST merece ser mais investigado, uma vez que esta desacetilase vem sendo sugerida como alvo potencial para o desenvolvimento de fármacos. Em conclusão, a atividade quimiopreventiva de EST e da TB envolve na hepatocarcinogênese experimental mecanismos epigenéticos que podem ou não ser distintos. / Hepatocellular carcinoma (HCC) has a poor prognosis, which makes its chemoprevention important. Tributyrin (TB), which is a histone deacetylase inhibitor (HDACi), is a promising chemopreventive agent of hepatocarcinogenesis. The chemopreventive effects of structured lipids (STLs) that were obtained by the enzymatic interesterification of TB with flaxseed oil (FSO) were evaluated in the present study. Rats were treated with STLs (STL group, 165 mg/100 g body weight (bw)), TB (TB group, 200 mg/100 g bw), FSO (FSO group, 133 mg/100 g bw), TB mixed with FSO (BLD group, 165 mg/100g bw) or maltodextrin (MD) (MD group; isocaloric control; 300 mg/100 g bw) daily for eight consecutive weeks by gavage. Two weeks after the initiation of treatment, the animals were subjected to the resistant hepatocyte hepatocarcinogenesis model (RH). The STL and TB groups developed blocker and suppressive chemopreventive activity against hepatocarcinogenesis, respectively. TB treatment induced apoptosis, unlike the STL treatment. Additionally, TB treatment resulted in the acetylation and trimethylation of H3K9 and H3K27, whereas the STLs acted only in the trimethylation of these histones. When analyzing the expression of genes involved in histone modifications, the STLs and TB reduced enhancer of zeste homolog 2 (Ezh2) and histone deacetylase 4 (Hdac4) gene expression. Conversely, only the STLs increased Hdac6 gene expression. This effect of the STLs warrants further investigation because this deacetylase has been suggested as a potential drug development target. In conclusion, the chemopreventive activities of the STLs and TB in experimental hepatocarcinogenesis involve epigenetic mechanisms that may be distinct.

A utilização da radioterapia e da quimioterapia em maior escala no tratamento do câncer de cabeça e pescoço tem elevado a incidência de efeitos colaterais, em especial da mucosite bucal. O presente estudo estabeleceu como objetivo a avaliação do efeito da suplementação da vitamina E e do selênio na prevenção da mucosite causada pela radioterapia e/ou quimioterapia em pacientes com neoplasias malignas das vias aerodigestivas superiores. Como metodologia, decidiu-se por um ensaio clínico III, randomizado, duplo cego, realizado no Serviço de Oncologia da Faculdade de Odontologia da Universidade Federal de Minas Gerais. A amostra foi constituída por 95 pacientes portadores de câncer nas vias aerodigestivas superiores, com indicação de radioterapia, sendo 78 (82,1%) do gênero masculino e 17 (17,9%) do gênero feminino, com média de idade de 54 ± 13,4 anos, variando entre 20 e 85 anos. O mineral selênio e a vitamina E foram classificados como grupo A e grupo B, respectivamente. Do primeiro dia da sessão de radioterapia até uma semana após o término da radioterapia, os pacientes tomaram uma cápsula por dia do suplemento. A mucosite foi avaliada semanalmente pelos cirurgiões dentistas do próprio serviço. Foram coletados dados de 115 pacientes. Como 20 deles foram excluídos, analisaram-se os resultados de apenas 95 indivíduos. Durante as dez semanas de acompanhamento desses pacientes, a avaliação da presença de mucosite foi realizada semanalmente. Do total de pacientes analisados, 88,2% desenvolveram mucosite (41 pacientes do grupo selênio e 41 pacientes do grupo vitamina E) decorrente do tratamento radioterápico realizado concomitantemente ou não com a quimioterapia. Na avaliação dos efeitos da suplementação nos diferentes graus de mucosite, observou-se que a maioria dos indivíduos se enquadrava na classificação grau II (alimentando-se normalmente, mas com dor). Independentemente do grau de mucosite, não foram encontradas diferenças entre os grupos suplementados (p=0,559). Em um período prévio a este trabalho, observou- se em pacientes avaliados semanalmente no mesmo ambulatório frequência de mucosite de 93% (noventa e três), dos quais 10,3% (dez) classificaram- se entre grau 0 e grau 1; 72,5% (setenta e dois) grau 2 e 17,2%.(dezessete) entre grau 3 e grau 4.Esses resultados sugerem que a suplementação com 100 µg de selênio ou de 400 mg de vitamina E não foi eficiente na prevenção da mucosite. Como conclusão, este estudo demonstra que a frequência de mucosite decorrente de tratamento radioterápico, com ou sem quimioterapia, é alta (88,2%) e que a suplementação do mineral selênio não é diferente da vitamina E na prevenção, intensidade ou retardo do aparecimento dessa enfermidade. / The increase of radiotherapy and chemotherapy intensity in the treatment of head and neck cancer has raised the incidence of side effects, specially of mouth mucositis. To evaluate the effect of the vitamin E and selenium supplementation in the prevention of the mucositis caused by the radiotherapy and/or chemotherapy in patients with upper aerodigestive ducts malign tumors. This study referred to at the Oncology Service of the UFMGs Oncology College. The sample has been constituted by patients with head and neck cancer and radiotherapy. 95 individuals were included, 78 (82,1%) male and 17 (17,9%) female, with a mean of age of 54+- 13,4, varying between 18 and 82 years old. The Selenium mineral and the Vitamin E were classified as Group A and B, respectively. The patients took one supplement capsule a day. The treatment started in the day of the radiotherapy session and ended one week after the radiotherapy . The mucositis was evaluated weekly by the services dentist surgeries. Data from 115 patients were collected. However, 20 patients were excluded from the study, therefore, 95 patients were analyzed. During the 10 weeks of patient follow-up, the mucositis presence evaluation was made weekly. We observed that 41 patients developed mucusitis during the radiotherapy treatment with or without chemotherapy, in both selenium and vitamin E groups. By evaluating the supplementation effects in the different levels of mucositis, it was observed that the great majority of individuals can be put in the level II classification (feed themselves normally, but with pain). Regardless the mucositis level, we have not found differences between the supplemented groups (p=0,559).In a period prior to this work, we observed in patients evaluated in the same clinic on a weekly frequency of mucositis 93% (ninety- three), of which 10.3% (ten) rated between grade 0 and grade 1, 72 , 5% (seventy two) grade 2 and 17.2%. (seventeen) of grade 3 and grade 4.We can suggest, therefore, that the supplementation with 10µg of selenium or 400mg of vitamin E have not been efficient in the mucositis prevention.In conclusion, this study has shown that the Selenium mineral and the Vitamin E supplementation did not prevent or retard the mucositis appearance.

Philosophiae Doctor - PhD / Fusarium verticil/ioides (Sacc) Nirenberg a common phytopathogen of maize and maize-based products produces fumonisin B (FB) mycotoxins that have been related to several diseases such as equine leukoencephalomalacia (ELEM), porcine pulmonary edema (PPE), liver toxicity in several animals and esophageal and liver cancer in humans. In one of our studies we hypothesize that aqueous extracts of indigenous South African wild garlic species (Tulbaghia violacea, T. alliacea and T. simmleri) may enhance the efficacy of the fungicides, SporekilPu, Thiram, Itraconazole and Fluconazole against F. verticil/ioides (MRC 826). Data analysis from in vitro results indicates that for the 16 different mixtures of each plant extract and fungicide combination, several significantly (P<O.05) higher growth inhibition responses were produced. More synergistic interactions were observed for the combinations of sporekill with T. violacea (62%) and T. alliacea (75%) than for T. simmleri (25%) .. Mixtures between the azole fungicides and T. simmleri produced 94 % synergistic interactions. Combination of fungicides and plant compounds offers the opportunity to find synergistic mixtures and may validate disease control strategies with increased biological activity and low dose rate application. Modulation studies of hepatic drug metabolizing enzymes and oxidative properties of AI/ium sativum, Tulbaghia violacea and T. alliacea in male Fischer rats were also evaluated. Due to its complex phytochemical composition a battery of assays were used to evaluate antioxidant potential. The extracts exhibited no adverse effects in the liver and kidneys of the rats. Total plasma iron was not affected showing no evidence for iron catalyzed lipid peroxidation. An increase was noted in hepatic ORAC values for rats consuming T. violacea and T. al/iacea. However, no correlation was observed between the phenolic intake by the rats and the increased hepatic ORAC levels. In this study, pre-treatment with aqueous extracts of T. violacea, T. al/iacea and A. sativum resulted in a significant elevation in GSH levels, induction of GST -IJ and UDP-GT and modulation of CAT and SOD. This modulated oxidative status and phase II drug metabolizing enzymes in the liver may protect the liver against the adverse effects related to oxidative damage and mutagenesis. The chemoprotective properties of crude aqueous extracts of A. sativum, T. violacea and T. alliacea were investigated on preneoplastic foci formation promoted by culture material of F. verticil/ioides MRC 826 utilizing diethylnitrosamine (DEN) as cancer initiator. Clinical chemical parameters related to liver and kidney function and decreased body weight gain suggesting that severe, acute liver injury had been induced in the positive control (DEN-eMF) rats, while the levels were mostly reduced by the garlic treatments. This study further indicates that T. alliaceae (2 % w/v) and A. sativum (1% w/v) treatment suppressed GST-P+ foci formation with the modulation of GST-0 phase II detoxification enzymes, as well as the antioxidant enzyme, SOD (T. alliaceae) and decreased GSH levels as being possible mechanisms of protection. These results provide new evidence showing the modulation of phase II drug metabolizing enzymes and the oxidative status in the liver of rats by the wild garlic species as well as A. sativum.

Indiana University-Purdue University Indianapolis (IUPUI) / The proteasome is responsible for breaking down the majority of the proteins in the cell. However, a complete understanding of how this large multi-subunit protease is assembled is currently lacking. Proper and timely assembly of the proteasome is critical for the functioning of the ubiquitin-proteasome pathway, defects in which have been associated with several different cancers. A recently discovered heterodimeric proteasome assembly chaperone, Pba1p-Pba2p, has been suggested to prevent the assembly process from straying off path. Pba1p-Pba2p associates with proteasomal assembly intermediates via C-terminal HbYX motifs. The HbYX motif is a tri-peptide sequence containing a hydrophobic residue (Hb) followed by a tyrosine (Y), then any amino acid (X). This motif was originally identified in proteasomal activators, and shown to mediate the association of activators with the proteasome by inserting into intersubunit pockets on either end of the proteasome. There are seven unique intersubunit binding pockets, located between neighboring α subunits on the proteasome, to which a HbYX-containing protein can bind; which of these pockets Pba1p-Pba2p binds to remains elusive. I attempted to identify where Pba1p and Pba2p bind via a crosslinking approach. Specific residues were mutagenized to cysteines on Pba1p, Pba2p, and the individual α subunits in order to generate crosslinkable species. By exposing yeast cells expressing these crosslinkable proteins to mild oxidizing conditions, I attempted to trap the Pba1p and Pba2p α intersubunit pocket interactions. In order to optimize crosslinking conditions, the assay was modified several ways. Additionally, measures were taken to increase detection of the crosslinked species via immunoblotting. Despite the efforts to improve the crosslinking and detection, I was unable to successfully detect a crosslinked species. However, crosslinking is a reasonable method to identify the Pba1p and Pba2p proteasomal binding sites, having been successfully used to identify binding sites for other HbYX-motif-containing proteins; further assay optimization should yield Pba1p and Pba2p proteasomal crosslinks.

No description available.

L'inflammation est considérée comme un promoteur de la cancérogenèse. La cyclooxygénase-2 (COX-2), la forme inductible de la famille des cyclooxygénases est un médiateur important de l'inflammation. Cette enzyme est constitutivement exprimée dans un grand nombre de cancers tels que les cancers du sein, du colon ou de la prostate. De nombreuses études mettent en évidence que COX-2 est surexprimée lors des étapes pré-néoplasiques. La COX-2 représente de ce fait une cible thérapeutique potentielle en chimioprévention et également pour le traitement des cancers. L'utilisation d'inhibiteurs synthétiques de COX-2 qui ciblent l'activité enzymatique est le seul traitement clinique actuellement disponible pour réduire l'activité de COX-2. Cependant, ces agents présentent des effets secondaires sévères, ce qui limite leur prise chronique chimiopréventives ou au cours des traitements anti-cancéreux. Une stratégie alternative pour cibler la fonction de COX-2 est d'inhiber son expression. Un grand nombre d'études montrent que certains produits naturels (la curcumine, le resveratrol ou l'apigénine par exemple) inhibent, préférentiellement l'expression de COX-2, sans être toxique. Notre projet analyse l'effet de l'acide cinnamique, un produit naturel extrait de la plante Cinnamonium cassia, sur l'expression de COX-2 au cours de la cancérogenèse dans le but d'évaluer son intérêt en chimioprévention. Nous avons utilisé comme modèle les cellules mammaires non carcinogènes, MCF10A stimulées avec un ester de phorbol, le 12-phorbol myristate 13-acetate (PMA), qui induit l'expression de COX-2. Nous avons observé une diminution de l'expression de COX-2 au niveau de l'ARNm et de la protéine après le traitement avec différentes concentrations d'acide cinnamique (1 et 10μM). L'analyse des mécanismes impliqués dans la diminution de l'expression de COX-2 a mis en évidence que l'acide cinnamique régule l'expression de COX-2 de façon post-transcriptionnelle en réduisant la stabilité de son ARNm. Cet effet est associé à une prévention de la diminution de l'expression de microARN (miR)-16 et une inhibition de l'expression de p38 induite par l'acide cinnamique en réponse au traitement avec le PMA. / Inflammation is considered a cancer-promoting factor. Cyclooxygenase-2 (COX-2), the inducible form of the family of cyclooxygenases is an important mediator of inflammation, which has been found constitutively expressed in many forms of cancer including breast, colon or prostate. A number of studies show that COX-2 is stably expressed since the early pre-neoplastic stages. This encourages us to consider COX-2 as a potential target in chemoprevention as well as in the treatment of cancer. Synthetic inhibitors of COX-2, which target enzymatic activity, are the only clinical strategy to counteract COX-2. However, these compounds present severe side effects, a fact that limits their prolonged intake, like requested in chemoprevention or during anti-cancer treatment.An alternative strategy to target COX-2 is at the level of its expression. A number of studies show that several natural compounds including curcumin, resveratrol or apigenin preferentially target COX-2 expression without showing toxicity.Our study analyses the effect of cinnamic acid, a natural compound derived from Cinnamonium cassia on COX-2 expression during carcinogenesis, with the final aim to evaluate its potential in chemoprevention/therapy.For our chemopreventive purposes, we used the non-carcinogenic breast cell line MCF10A, stimulated by the phorbol ester 12-phorbol myristate 13-acetate (PMA), which typically induces COX-2. We show a reduction of induced COX-2 expression after treatment with different concentrations of cinnamic acid (1 and 10μM). The analysis of the mechanisms involved in COX-2 protein expression decrease shows that cinnamic acid regulated COX-2 expression at the post-transcriptional level by reducing COX-2 mRNA stability. This effect is associated with the ability of cinnamic acid to prevent downregulation of miR-16 expression and p38 activation in response to PMA treatment.

A combinação de agentes quimiopreventivos com diferentes mecanismos de ação tem sido considerada uma estratégia promissora para a prevenção do câncer. Dentre os diversos compostos bioativos em alimentos, destacam-se a tributirina, um pró-fármaco do ácido butírico presente em laticínios e produzido pela fermentação de fibras dietéticas, e o óleo de linhaça, fonte de ácido alfa linolênico. Nesse contexto, foi avaliada a atividade quimiopreventiva de lipídios estruturados obtidos a partir da interesterificação enzimática de tributirina e óleo de linhaça durante a fase de promoção inicial da hepatocarcinogênese experimental. Ratos Wistar machos submetidos ao modelo do hepatócito resistente receberam diariamente, por via intragástrica (i.g), maltodextrina, óleo de linhaça, tributirina, a mistura não esterificada ou lipídios estruturados durante a fase de promoção inicial. O tratamento com lipídios estruturados demonstrou atividade quimiopreventiva comparável à da tributirina, mesmo resultando em menor concentração hepática de ácido butírico. Tanto a tributirina quanto os lipídios estruturados não inibiram a proliferação celular em lesões preneoplásicas, mas induziram a apoptose naquelas em remodelação. Os efeitos inibitórios da tributirina em fases iniciais da hepatocarcinogênese experimental estão relacionados ao aumento da acetilação de histonas e à modulação de processos de translocação nuclear da p53. No presente estudo, foi observado aumento substancial da razão nuclear/citoplasmática de p53 e importina-alfa em fígados de animais submetidos ao modelo e tratados com tributirina, mas não nos tratados com lipídios estruturados. Por outro lado, o tratamento com lipídios estruturados reduziu a expressão dos oncogenes Bcl2, Ccnd2, Pdgfa, Vegfa e aumentou a expressão dos genes supressores de tumor Cdh13, Fhit e Socs3. Assim, embora o potencial quimiopreventivo dos lipídios estruturados seja comparável ao da tributirina, os resultados sugerem que o novo composto não exibe atividade de HDACi, e que seus efeitos inibitórios na hepatocarcinogênese possam ser atribuídos à modulação da expressão de oncogenes e genes supressores de tumor. / Combination of chemopreventive agents with different mechanisms of action has been considered a promising strategy to cancer prevention. Among several bioactive food compounds, tributyrin, a butyric acid prodrug obtained from dairy products and dietetic fiber fermentation, and flax seed oil, a rich source of alpha linolenic acid have shown chemopreventive potential. Here, we evaluated the chemopreventive activity of structured lipids obtained by enzymatic interesterification of tributyrin and flax seed oil during the early promotion phase of experimental hepatocarcinogenesis. Male Wistar rats subjected to the resistant hepatocyte model were treated daily, i.g, with maltodextrin, flax seed oil, tributyrin, non-sterified blend, or structured lipids. Treatment structured lipids showed similar chemopreventive activity compared to tributyrin, even when structured lipids yielded lower concentrations of butyric in the liver. Tributyrin and structured lipids did not inhibit cell proliferation in preneoplastic lesions, but both of them induced apoptosis in remodeling preneoplastic lesions. In addition, histone acetylation and p21 restored expression tributyrin molecular mechanisms were related to modulation of p53 nuclear shuttling mechanisms. In the present study, it was observed a substantial increase in p53 nuclear/cytoplasmic ratio and importin-alpha in preneoplastic livers of tributyrin treated rats, but not in those treated with structured lipids. In contrast, treatment structured lipids downregulated expression of major oncogenes Bcl2, Ccnd2, Pdgfa, and Vegfa; and upregulated expression of critical tumor suppressor genes, Cdh13, Socs3 and Fhit. Hence, although structured lipids and tributyrin show similar chemopreventive potential, the results suggest that the new compound does not exhibit HDACi activity, and that its inhibitory effects may be attributed to the modulation of oncogenes and tumor suppressor genes expression.

La contaminazione da micotossine può coinvolgere tutti i comparti della filiera agroalimentare e rappresenta a tutti gli effetti un potenziale di perdita economica. Il settore zootecnico è particolarmente esposto alle ricadute della contaminazione, in termini di costi sanitari per gli effetti cronici sulla salute animale conseguenti all’esposizione alle micotossine. La prevenzione in campo mediata da appropriate tecniche agronomiche rappresenta, per quanto ovvio, la più importante strategia per ridurre la contaminazione dei mangimi. Un approccio completamente differente è quello che si basa sulla possibilità di ridurre gli effetti delle tossine sull’animale, modificando la sua alimentazione attraverso l’integrazione nutrizionale di opportuni agenti chemiopreventivi. Numerose sono le specie chimiche, di origine naturale o sintetica, che hanno dimostrato una efficacia chemiopreventiva del danno ossidativo indotto da micotossine ed emergono per la loro efficacia il selenio e i suoi composti organici ed inorganici. L’azione antineoplastica del selenio è nota da molto tempo e ha trovato conferme in numerosi studi epidemiologici che indicano una relazione inversa tra assunzione di selenio con la dieta e rischio di sviluppo di una patologia neoplastica. Questa tesi di dottorato ha avuto come obiettivo la progettazione, la sintesi e lo studio di composti diamminoacidici contenenti un atomo di calcogeno (selenio o zolfo), mimici dalla glutatione perossidasi (GPx) e auspicabilmente attivi nel contrasto del danno ossidativo indotto dall’aflatossina B1 (AFB1). Le specie diamminoacidiche sintetizzate appartengono alla classe dei Sec-derivati, ovvero sono costitute da un residuo di L-selenocisteina (o L-cisteina) a cui è legato attraverso l’atomo di calcogeno un altro L-amminoacido. Alla porzione calcogenica, che rappresenta il sito redox, sono stati infatti ancorati rispettivamente un residuo di L-prolina modificata e un residuo di L-leucina. Al termine delle fasi di sintesi e caratterizzazione chimico-fisica dei calco-diamminoacidi è stata intrapresa una serie di indagini biochimiche in collaborazione con due distinti laboratori in Germania. I risultati ottenuti hanno permesso di dimostrare che alcuni Sec-derivati sono in grado di ridurre i perossidi attraverso un meccanismo catalitico che mima l’attività della GPx. Essi sembrano inoltre evidenziare che il “meccanismo antiossidante” sia mediato sostanzialmente dall’attività GPx mentre non sono particolarmente attivi meccanismi di radical scavenging né di puro trasferimento elettronico. Alcuni dei Sec-derivati sintetizzati sono stati oggetto di un’indagine cellulare preliminare volta a verificare una eventuale effettiva azione protettiva dei calco-diamminoacidi verso il danno cellulare indotto da AFB1. Le indagini sono state effettuate adoperando una linea cellulare HepG2. I dati ottenuti, nel loro insieme, hanno mostrato che i calco-diamminoacidi esaminati sono di per sé caratterizzati da una notevole attività biologica. In particolare, alla concentrazione più elevata utilizzata (100 µM), alcuni di essi hanno mostrato interessanti proprietà citoprotettive contro il danno indotto da AFB1. L’efficacia è paragonabile, e in qualche caso superiore, a quella riscontrabile per il composto di riferimento, la Se-metil-selenocisteina, principale forma di selenio organico che è naturalmente presente in specie vegetali quali Allium Sativum e Brassica juncea. / Mycotoxin contamination may involve all fields of the food agricultural chain and might potentially determine economic losses. In particular, the livestock sector is at risk of health costs, as a consequence of chronic diseases induced by mycotoxin exposure. It is obvious, although actual, that the mycotoxin contamination should be controlled on the field by a series of suitable agriculture practices in order to minimize the mycotoxin formation in feeds. Recently, dietary strategies (antioxidant compounds, medicinal herbs, plant extracts) to counteract the effects of mycotoxins have attracted considerable attention. Chemoprevention refers to the use of naturally occuring and/or synthetic chemicals to inhibit the development of mycotoxin induced chronic diseases. Selenium and its organic and inorganic compounds, have emerged as outstanding chemoprevention agents. Selenium antineoplastic properties have been well documented through the last decades, being supported by several epidemiological evidences on the association between selenium intake and the risk for some kinds of human/animal cancers. This PhD thesis was focussed on design, synthesis and study of a family of chalcogenic diamino acids (containing either selenium or sulfur), GPx mimics and potentially active to reduce the oxidative damage induced by aflatoxin B1 (AFB1). The diamino acids synthesized belong to the family of Sec-derivatives and consist of L-selenocysteine (or L-cysteine) linked through the calchogen atom to either a L-proline or a L-leucine moiety. The GPx-like catalytic activity of the various compounds was confirmed using both thiophenol and GPx assays. Their possible radical-scavenging activity, and electron-donor reducing character as well, could be ruled out by suitable assays. The cytoprotective capacity exhibited by several of the compounds synthesized was investigated in HepG2 cells by MTS assays. The results indicated that all the chalcogenic diamino acids tested had a strong biological activity. In particular, the pre-treatment of the test cells with two Sec-derivatives, employed at higher doses (100 µM), led to a protective effect on cell toxicity, lowering the AFB1-induced cell mortality, and this efficacy is comparable with that of the reference Se-metil-selenocysteine, main source of organic selenium in plants such as Allium Sativum and Brassica juncea.