Made available in DSpace on 2014-12-17T14:03:40Z (GMT). No. of bitstreams: 1 LuizaSEPW_DISSERT.pdf: 2731256 bytes, checksum: a154afda557a95d39d943b76672810db (MD5) Previous issue date: 2012-12-21 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / This study examines the physical and chemical composition and the pharmacological effects of brown seaweed FRF 0.8 Lobophora variegata. Fractionation of the crude extract was done with the concentration of 0.8 volumes of acetone, obtaining the FRF 0.8. The physicochemical characterization showed that it was a fucana sulfated. Anti-inflammatory activity was assessed by paw edema model by the high rates of inhibition of the edema and the best results were in the fourth hour after induction (100 ? 1.4% at the dose of 75 mg / kg) and by the strong inhibitory activity of the enzyme myeloperoxidase (91.45% at the dose of 25 mg / kg). The hepataprote??o was demonstrated by measurements of enzymatic and metabolic parameters indicative of liver damage, such as bilirubin (reduction in 68.81%, 70.68% and 68.21% for bilirubin total, direct and indirect, respectively at a dose of 75 mg / kg), ALT, AST and γ-GT (decrease of 76.93%, 44.58% and 50% respectively at a dose of 75 mg / kg) by analysis of histological slides of liver tissue, confirming that hepatoprotective effect the polymers of carbohydrates, showing a reduction in tissue damage caused by CCl4 and the inhibition of the enzyme complex of cytochrome P 450 (increasing sleep time in 54.6% and reducing the latency time in 71.43%). The effectiveness of the FRF 0.8 angiogenesis was examined in chorioallantoic membrane (CAM) of fertilized eggs, with the density of capillaries evaluated and scored, showing an effect proangig?nico at all concentrations tested FRF (10 mg- 1000 mg). The FRF showed antioxidant activity on free radicals (by inhibiting Superoxide Radical in 55.62 ? 2.10%, Lipid Peroxidation in 100.15 ? 0.01%, Hydroxyl Radical in 41.84 ? 0.001% and 71.47 Peroxide in ? 2.69% at concentration of 0.62 mg / mL). The anticoagulant activity was observed with prolongation of activated partial thromboplastin time (aPTT) at 50 mg (> 240 s), showing that its action occurs in the intrinsic pathway of the coagulation cascade. Thus, our results indicate that these sulfated polysaccharides are an important pharmacological target / Este estudo analisa a composi??o f?sico-qu?mica e os efeitos farmacol?gicos da FRF 0,8 da alga marrom Lobophora variegata. O fracionamento do extrato bruto foi feito com a concentra??o de 0,8 volumes de acetona, obtendo a FRF 0,8. A caracteriza??o f?sico-qu?mica mostrou que se tratava de uma fucana sulfatada. Foi verificada a atividade antinflamat?ria pelo modelo de edema de pata, atrav?s das altas taxas de inibi??o do edema e os melhores resultados foram na quarta hora ap?s a indu??o (100 ? 1,4% com a dose de 75 mg/kg) e pela forte atividade inibidora da enzima mieloperoxidase (91,45% com a dose de 25 mg/kg). A hepataprote??o foi demonstrada pelas dosagens de par?metros metab?licos e enzim?ticos indicativos de dano hep?tico, como bilirrubina (redu??o em 68,81%, 70,68% e 68,21% para bilirrubinas total, direta e indireta, respectivamente na dose de 75 mg/kg), ALT , AST e γ-GT (diminui??o em 76,93%, 44,58% e 50%, respectivamente na dose de 75 mg/kg), pela an?lise das l?minas histol?gicas do tecido hep?tico, que confirmam esse efeito hepatoprotetor dos pol?meros de carboidrato, mostrando uma redu??o no dano tecidual causado por CCl4, e pela inibi??o do complexo enzim?tico do citocromo P 450 (aumentando o tempo de sono em 54,6% e reduzindo o tempo de lat?ncia em 71,43%). A efic?cia sobre a angiogenese da FRF 0,8 foi examinada na membrana corioalant?ica (CAM) de ovos fertilizados, com a densidade dos capilares avaliadas e pontuadas, mostrando um efeito proangig?nico em todas as concentra??es de FRF testadas (10 μg-1000 μg). A FRF apresentou a??o antioxidante sobre radicais livres (inibindo o Radical Super?xido em 55,62?2,10%, Peroxida??o Lip?dica em 100,15?0,01%, Radical Hidroxila em 41,84?0,001% e Per?xido em 71,47?2,69%, todos na concentra??o de 0,62 mg/mL). A atividade anticoagulante foi verificada com o prolongamento do tempo de tromboplastina parcial ativada (aPTT) a 50 μg (>240 s), mostrando que sua a??o ocorre na via intr?nseca da cascata de coagula??o. Sendo assim, nossos resultados indicam que estes polissacar?deos sulfatados constituem um importante alvo farmacol?gico

Submitted by isabela.moljf@hotmail.com (isabela.moljf@hotmail.com) on 2017-05-12T13:37:04Z No. of bitstreams: 1 pedroluizrodriguesguedes.pdf: 3448334 bytes, checksum: 3bac263ee478fd60c02c5d6036a0e81e (MD5) / Approved for entry into archive by Adriana Oliveira (adriana.oliveira@ufjf.edu.br) on 2017-05-12T15:44:09Z (GMT) No. of bitstreams: 1 pedroluizrodriguesguedes.pdf: 3448334 bytes, checksum: 3bac263ee478fd60c02c5d6036a0e81e (MD5) / Made available in DSpace on 2017-05-12T15:44:09Z (GMT). No. of bitstreams: 1 pedroluizrodriguesguedes.pdf: 3448334 bytes, checksum: 3bac263ee478fd60c02c5d6036a0e81e (MD5) Previous issue date: 2013-07-30 / CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Condroitim sulfato (CS) é um glicosaminoglicano (GAG), presente na matriz extracelular (MEC) de vários tecidos de mamíferos, utilizado para o tratamento da osteoartrite e, recentemente, tem despertado grande interesse devido ao seu potencial anti-inflamatório. Vários modelos experimentais in vivo de inflamação são empregados para o estudo da atividade anti-inflamatória, entre eles o modelo de fibrose induzida por colestase extra-hepática. A colestase produz lesão hepatocelular com edema do trato portal, infiltrado leucocitário, proliferação de células epiteliais biliares e fibrose do trato portal. O objetivo deste trabalho foi analisar os efeitos do CS no modelo de colestase extra-hepática emperimental induzido por laqueadura do ducto biliar (BDL) em ratos Wistar. Para isso foram utilizados animais (n = 82) de 6 a 8 semanas de idade eutanasiados 2, 7 ou 14 dias após o procedimento cirúrgico divididos nos grupos: BDL, BDL tratado com CS, Sham e Sham tratado com CS. Foram avaliados peso corporal e do fígado dos animais, concentrações séricas de bilirrubina direta (BD), globulinas, atividades de gama glutamil transpeptidase (Gama GT), fosfatase alcalina (FA), alanina transaminase (ALT) e aspartato transaminase (AST), alterações morfológicas no tecido, atividade de mieloperoxidase (MPO), atividade de metaloproteases (MMP-9, MMP-2 e pró MMP-2) e conteúdo de GAGs no fígado dos animais, além da análise histopatológica do tecido hepático. O CS obtido para a realização do trabalho apresentou teores superiores a 92%, com peso molecular de aproximadamente 40 kDa e um conteúdo dissacarídico com predominância de Δdi4S (65%). BDL gerou vários sintomas relacionados à lesão celular e ao processo inflamatório como aumento dos níveis séricos de BD e globulinas, aumento das atividades de Gama GT, FA, ALT e AST, infiltrado inflamatório e modificação morfométrica, com proliferação ductular, e na MEC do fígado dos animais induzidos. CS levou a redução do aumento inicial das transaminases indicando proteção dos tecidos lesados no procedimento cirúrgico. O tratamento levou à redução do infiltrado inflamatório no tecido, expresso pela diminuição significativa da atividade de MPO no homogenato. A remodelação tecidual também foi reduzida, havendo diminuição da atividade de MMP-9, pró MMP-2 e MMP-2 e ainda dos níveis dos GAGs dermatam sulfato e heparam sulfato presentes, produzidos por células estreladas em resposta ao dano no tecido. Estes resultados mostram que o CS reduziu os efeitos da lesão hepática do modelo e foi capaz de retardar a fibrogênese hepática. / Chondroitin sulfate (CS) is a glycosaminoglycan (GAG) present in the extracellular matrix (ECM) of many mammalian tissue, used for osteoarthritis treatment and, recently, has aroused great interest due to its anti-inflammatory potential. Several in vivo inflammation experimental models are employed to study anti-inflammatory activity, including extra-hepatic cholestasis induced fibrosis. Cholestasis produces hepatocellular injury with portal tract edema, leukocyte infiltration, biliary epithelial cells proliferation and portal tract fibrosis. The aim of this work was to analyze CS effects on an extra-hepatic cholestasis experimental model induced by bile duct ligation (BDL) on Wistar rats. For this purpose 6 to 8 weeks old animals (n = 82) were euthanized 2, 7 or 14 days after surgical procedure, previously divided into groups: BDL, CS treated BDL, Sham, CS treated Sham. To analyze disease evolution body and liver weight, serum concentrations of direct bilirubin (BD), globulins, activities of gamma glutamyl transferase (Gama GT), alkaline phosphatase (FA), alanine and aspartate aminotransferases (ALT and AST), morphological changes on tissue, mieloperoxidase (MPO) activity, matrix metalloproteinases (MMP-9, pró MMP-2 and MMP-2) activities and liver GAGs content, besides histopathological analysis of the tissue. CS acquired presented over 92% tenor, molecular weight of approximately 40 kDa and disaccharide content of Δdi4S predominantly (65%).BDL caused many symptoms related to cellular damage and inflammatory process such as increasing BD and globulins, elevation of Gama GT, FA, ALT and AST activities, inflammatory infiltrate and changes on liver morphometry, with ductular proliferation, and on the ECM. CS reduced initial burst on aminotransferases, indicating protection of tissues injured on surgery procedure. Treatment led to reduction of inflammatory infiltrate, showed by significant decreasing on MPO activity. Tissue remodeling was also reduced, with decrease of MMP-9, pro MMP-2 and MMP-2 activities and also of GAGs dermatam sulfate and heparam sulfate levels, produced by hepatic stellate cells in response of tissue damage. These results show that CS reduced cholestasis hepatic injury effects, being capable to slow down liver fibrogenesis.

Le carcinome hépatocellulaire (CHC) est un cancer au pronostic sombre, car il est souvent diagnostiqué trop tardivement pour entreprendre un traitement curatif. Il se développe dans 80-90% des cas sur fond de cirrhose. On connait mal comment la fibrose, étape préliminaire à la cirrhose, et son principal constituant, le collagène de type 1 (COL1), peuvent jouer un rôle dans le processus du CHC. Nous avons tout d’abord étudié le développement de la fibrose dans un modèle utilisant la souris nue. Nous avons déterminé qu’après 16 semaines d’administration de thioacétamide dans l’eau de boisson, il est possible d’obtenir une fibrose suffisante pour induire une hépatoprotection en présence de différents hépatotoxiques (AST dans le sérum de souris fibrotiques vs non-fibrotiques : Anti-Fas JO2 (4665 ± 2596 vs. 13953 ± 2260 U/L; P<0.05), acétaminophène (292 ± 66 vs. 4087 ± 2205 U/L; P<0.01) et CCL4 (888 ± 268 vs. 15673 ± 2782 U/L; P<0.001)). Ces résultats confirment que la présence de COL1 et de fibrose favorise la survie des hépatocytes normaux tel qu’observé précédemment au laboratoire. Par la suite, nous avons sélectionné in vivo, par injection intrasplénique de la lignée de CHC Hepa1-6, une lignée à forte tumorigénicité nommée dt-Hepa1-6 (28±12 lésions vs. 0±0 lésions à 21 jours). Cette lignée était composée d’une sous-population cellulaire arborant la protéine de surface EpCAM (34.0±0.1%). Par tri cellulaire, nous avons démontré que ces cellules étaient partiellement responsables de la tumorigénicité accrue (EpCAM + (86.7±2.3%) :1093±74 lésions vs. EpCAM- (15.3±1.0%) :473±100 lésions; P<0.01). Nous avons alors démontré que la présence de fibrose favorise le développement de la lignée dt-Hepa1-6 in vivo (604±242 vs 22±9 lésions; P<0.05). De plus, la présence de fibrose réduit l’efficacité du traitement au cisplatin in vivo (44.5±4.9 vs. 78.7±6.9%; P<0.01) confirmant les résultats obtenus in vitro (Apoptose : COL1 13.75±0.44% vs. plastique 31.45±1.37%; P<0.001). En conclusion, la présence de fibrose et de son principal constituant, le COL1, favorise la survie et la progression du CHC. / Hepatocellular carcinoma (HCC) is a dreadful pathology, often diagnosed too late to be cured. In 80-90% of cases, it arises in the context of liver cirrhosis. Little is known on the implication of liver fibrosis, one of the key elements of cirrhosis, and its major constituent, type I collagen (COL1), on the development of HCC. We first studied the development of fibrosis in a nude mouse model. We determined that, after 16 weeks of thioacetamide administration in drinking water, we obtained a sufficient degree of fibrosis to reach a hepatoprotective state when animals were exposed to different hepatotoxic agents (Serum AST of fibrotic vs non-fibrotic mice : Anti-Fas JO2 (4665 ± 2596 vs. 13953 ± 2260 U/L; P<0.05), acetaminophen (292 ± 66 vs. 4087 ± 2205 U/L; P<0.01) et CCL4 (888 ± 268 vs. 15673 ± 2782 U/L; P<0.001)). This confirmed that COL1 and the presence of fibrosis protects normal hepatocytes as observed previously in our laboratory. Next, we selected in vivo, by intrasplenic injection of the murine HCC cell line Hepa1-6, a highly tumorigenic cell line that we named dt-Hepa1-6 (28±12 lesions vs. 0±0 lesions at 21 days). This cell line was constituted of cell subsets expressing EpCAM protein at their surface (34.0±0.1%). Through cell sorting, we demonstrated that these cells were partially responsible for the enhanced tumorigenicity observed (EpCAM + (86.7±2.3%) :1093±74 lesions vs. EpCAM- (15.3±1.0%) :473±100 lesions; P<0.01). We then showed that the presence of liver fibrosis increases the development of dt-Hepa1-6 cell line in vivo (604±242 vs 22±9 lesions; P<0.05). Moreover, fibrosis reduced the anti-neoplastic efficacy of cisplatinum in vivo (44.5±4.9 vs. 78.7±6.9%; P<0.01) confirming in vitro results (Apototic index : COL1 13.75±0.44% vs. plastic 31.45±1.37%; P<0.001). In conclusion, fibrosis and its major constituent, COL1, favor the survival and progression of HCC.

Background & Aims: Oxidative stress and apoptosis are proposed mechanisms of cellular injury in studies of xenobiotic hepatotoxicity. The aim of this work is to find early signal markers of drug-induced injury of the liver by focusing on select antioxidant/oxidant and apoptotic genes. As well, to address the relationship between conventional liver dysfunction markers and the measured mRNA and protein expressions in the D-galactosamine/lipopolysaccharide and tert-butylhydroperoxide hepatotoxicity models. Furthermore, potential hepatoprotective capabilities of antioxidant polyphenols quercetin and curcumin were evaluated in relation to its modulation of the oxidative stress and apoptotic parameters in the given xenobiotic hepatotoxicity models. Methods: Biochemical markers testing the hepatic function included aminotransferases (ALT, AST) and bilirubin. Measurements of TBARS and conjugated dienes were used to assess lipoperoxidation. Plasma levels of catalase and reduced glutathione were used as indicators of the oxidative status of the cell. Real time PCR was used to analyse the mRNA expressions of the inducible nitric oxide synthase (NOS-2), heme oxygenase-1 (HO-1), superoxide dismutase (SOD-1), glutathione peroxidase (Gpx-1), caspase 3 (Casp3), BH3 interacting domain death agonist (Bid) and Bcl-2...