Submitted by Marcos Anselmo (marcos.anselmo@unipampa.edu.br) on 2016-04-04T17:31:47Z No. of bitstreams: 1 Marcelo Gomes de Gomes.pdf: 1949846 bytes, checksum: c4472edd9df88ec1d4f0e86bd59fafa9 (MD5) / Made available in DSpace on 2016-04-04T17:31:47Z (GMT). No. of bitstreams: 1 Marcelo Gomes de Gomes.pdf: 1949846 bytes, checksum: c4472edd9df88ec1d4f0e86bd59fafa9 (MD5) Previous issue date: 2015-03-02 / O lipopolissacarídeo (LPS) associado com a D-galactosamina (D-GalN), tetracloreto de carbono (CCl4) e o paracetamol (PCM), são modelos animais de hepatite fulminante aguda amplamente utilizados. O gama orizanol (γ-ORY) é uma mistura de 10 ésteres de ácido trans ferúlico e é encontrado no óleo do farelo de arroz. O objetivo deste estudo foi avaliar o efeito hepatoprotetor do γ-ORY nos diferentes modelos animais de hepatite fulminante aguda em camundongos. O experimento foi realizado através do tratamento com γ-ORY (50 mg/kg, por via oral, p.o.) durante uma semana, após uma hora do último tratamento foi administrado por via intraperitoneal o LPS (50 mg/kg) e D-GalN (500 mg/kg), foi utilizado silimarina como controle positivo. O γ-ORY foi administrado oralmente (50 mg/kg) durante uma semana, após 1 hora da última administração o dano foi induzido por uma única dose de CCI4 (1 mg/kg i.p.), a silimarina foi utilizada como controle positivo. Os animais receberam γ-ORY durante dois dias em duas doses (10 mg e 50 mg/kg p.o.), no terceiro dia o PCM (2 g/kg, p.o.). Após 24 horas os animais foram submetidos à eutanásia. Foram feitas as seguintes determinações bioquímicas: marcadores de estresse oxidativo (TBARS, NPSH, 4-HNE), defesas antioxidantes enzimáticas (CAT, SOD, GPx), marcadores de dano hepático (AST, ALT, GGT), marcadores de funcionalidade hepática (bilirrubina), marcadores inflamatórios (TNF-α, IL-1β), marcadores de apoptoses (Caspase 3 e 9) e análise histológica. O γ-ORY foi capaz de proteger significantemente em todas as determinações analisadas, podendo ser comparado ao um fármaco utilizado para o tratamento da hepatite fulminante aguda que é a silimarina. Portanto, sugerimos que o γ-ORY pode ser uma alternativa para o tratamento da hepatite fulminante aguda, pois ficou comprovado seu efeito hepatoprotetor está associado com sua capacidade antioxidante, anti-inflamatória e anti-apoptótica. / The lipopolysaccharide (LPS) associated with D-galactosamine (D-GalN), carbon tetrachloride (CCl4) and paracetamol (PCM) are acute fulminant hepatitis widely used animal models. The gamma oryzanol (γ-ORY) is a mixture of 10 esters of trans-ferulic acid and is found in rice bran oil. The objective was to assess the potential hepatoprotective effect of γ-ORY on mechanisms involved in different animal models of acute fulminant hepatitis in mice. The animals were divided into different groups depending on the experiment. γ-ORY treatment (50 mg/kg, per oral, p.o.) for a week after an hour of the last treatment before the administration by intraperitoneal of LPS (50 mg/kg) and D-GalN (500 mg/kg). γ-ORY was administered orally (50 mg/kg) for one week, after 1 hour of the last administration damage was induced by a single dose of CCl4 (1 mg /kg i.p.). The animals received γ-ORY for two days in two doses (10mg and 50mg /kg p.o.), on the third day PCM (2 g/ kg, p.o.). After 24 hours the animals were euthanized. The following biochemical determinations were made: non-enzymatic antioxidant defenses, defenses enzymatic antioxidants, liver damage markers, liver function markers, inflammatory markers, apoptosis markers and histological analysis. The γ-ORY was able to significantly protect in all analyzed determinations, and can be compared to a drug used for acute fulminant hepatitis. Therefore, we suggest that the γ-ORY can be an alternative for the treatment of acute fulminant hepatitis, as was proved its hepatoprotective effect is associated with its antioxidant capacity, anti-inflammatory and anti-apoptotic effect.

Introdução: O envolvimento hepático na doença celíaca (DC) é amplamente reconhecido e atualmente é uma das manifestações extra-intestinais mais freqüentes. Com o advento de marcadores sorológicos de elevada especificidade e sensibilidade, sobretudo o anticorpo antiendomísio (EMA), a DC tem sido descrita em associação a várias hepatopatias. Objetivos: caracterizar as formas de triagem de DC em portadores de hepatopatia crônica; caracterizar e estudar os pacientes cujo diagnóstico de DC foi realizado durante a investigação de uma doença hepática; pesquisar a reatividade do antiendomísio em pacientes com hepatite auto-imune, cirrose biliar primária, colangite esclerosante primária e hipertensão portal não cirrótica; avaliar o comportamento da doença hepática na vigência de dieta sem glúten. Métodos: Os pacientes foram triados pela detecção dos anticorpos anti-reticulina e anticorpo antimatriz extracelular durante a rotina de imunofluorescência de pesquisa dos auto-anticorpos hepáticos; pela presença de manifestações de DC em hepatopatas crônicos, pelo aspecto endoscópico sugestivo de DC e pela pesquisa sistemática do EMA nas patologias referidas anteriormente. Todos os pacientes foram submetidos à pesquisa do EMA, anti-reticulina IgG ou antimatriz de fibroblastos IgG na presença de deficiência de IgA. Em caso de positividade desses marcadores, foram submetidos à endoscopia digestiva alta para biópsia intestinal e caracterizados do ponto de vista clínico, laboratorial e histopatológico. A evolução desses dados permitiu a caracterização da evolução da doença hepática e da doença celíaca a partir da introdução da dieta sem glúten. Resultados: Foram identificados 43 pacientes com auto-anticorpos relacionados à DC (em 42 o EMA IgA e em um o antimatriz extracelular IgG em associação com deficiência de IgA). A rotina de pesquisa de auto-anticorpos hepáticos identificou 31 pacientes; seis apresentavam hepatopatia crônica e manifestação de DC; em três o exame endoscópico foi sugestivo de DC e a pesquisa sistemática do EMA foi positiva em três casos. O diagnóstico de DC foi confirmado em 37 de 40 pacientes (92,5%) em que a biópsia intestinal foi realizada. A idade dos pacientes variou de 2 a 68 anos, com mediana de 35 anos. Houve maior prevalência de acometimento no sexo feminino (65%). A DC foi mais prevalente na raça branca (87%), mais foi identificada em quatro mulatos e um negro. As doenças hepáticas mais freqüentes foram hipertransaminasemia criptogênica, hepatite auto-imune, hiperplasia nodular regenerativa e hepatite pelo vírus C. Conclusões: 1) A reatividade do anti-reticulina, a presença de diarréia inexplicada e a análise endoscópica da mucosa duodenal foram as formas de seleção mais efetivas de se identificar a DC em hepatopatas crônicos. 2) A ausência de manifestações clínicas de DC nesse grupo de pacientes foi bastante expressiva. 3) A pesquisa sistemática do EMA em cirrose biliar primaria, hepatite auto-imune, colangite esclerosante primária não contribuiu para o diagnóstico de DC em um número significativo de pacientes, ao contrário do observado no grupo de hipertensão portal não cirrótica, especialmente hiperplasia nodular regenerativa 4) As doenças hepáticas em que mais freqüentemente foi diagnosticada a DC foram a hiperplasia nodular regenerativa, hepatite auto-imune, hipertransaminasemia criptogênica, hepatite pelo vírus C e cirrose biliar primária antimitocôndria negativo. 5) A retirada do glúten da dieta contribuiu de maneira efetiva para normalização das enzimas hepáticas nos casos de hipertransaminasemia criptogênica. Nos grupos de hiperplasia nodular regenerativa, hepatite B e C, cirrose biliar primária, álcool e hepatite auto-imune, o papel da dieta foi de difícil avaliação em razão da interferência da instituição do tratamento específico e da evolução natural da doença hepática de base. / Introduction: The hepatic involvement in Celiac Disease (CD) is well known and widely regarded as a frequent extra-intestinal manifestation. With the advent of highly specific and sensitive serological markers, especially the antiendomysial antibody (EMA), CD has been described in association with several liver conditions. Objectives: To characterize ways for screening patients with chronic liver conditions in order to diagnose CD, to characterize and study patients whose CD diagnoses were performed when investigating hepatic diseases, to test the reactivity of EMA in patients with autoimmune hepatitis, primary biliary cirrhosis, primary sclerosing cholangitis and non-cirrhotic portal hypertension, to evaluate the course of the hepatic disease in gluten-free diet. Methods: Patients were selected by the detection of antireticulin and anti-extracellular matrix antibodies during routine immunoflourescence determination for hepatic autoantibodies, by the presence of CD manifestations in chronic patients with liver diseases, by the endoscopic aspects suggestive of CD and by systematic search for EMA in the above mentioned pathologies. All patients were submitted to tests for EMA, antireticulin IgG or antimatrix of IgG fibroblasts in IgA deficiency. When testing positive for these markers, patients were submitted to upper digestive endoscopy for intestinal biopsy, and were also characterized from the clinical, laboratorial and histological point of view. The assessment of these data enabled the characterization of the hepatic condition as well as the CD from the onset of a gluten-free diet. Results: 43 patients with autoantibodies related to CD were identified (42 tested positive for IgA EMA and 1 for IgG extracellular antimatrix in the presence of IgA deficiency). Routine determination of hepatic autoantibodies identified 31 patients. Of those, 6 presented chronic liver diseases and CD manifestations. In 3 patients, the endoscopic exam was suggestive of CD; systematic EMA determination was positive in all three cases. The CD diagnosis was confirmed in 37 out of 40 patients (92.5%) that performed intestinal biopsy. Patients aged between 2 and 68 years (median: 35 years). Female patients were most affected (65%). CD was more prevalent in white patients (87%), but was also found in four mulattoes and 1 black person. The most common liver disorders were cryptogenic hypertransaminasemia, autoimmune hepatitis, nodular regenerative hyperplasia and chronic hepatitis C. Conclusions: 1) The reactivity of antireticulin, the presence of unexplained diarrhea and the endoscopic analysis of the duodenal mucous were the most effective ways to identify CD in chronic liver diseases. 2) The absence of CD clinical manifestations in this group of patients was impressive. 3) Contrary to what was observed in the group with non-cirrhotic portal hypertension, especially regenerative nodular hyperplasia, the systematic determination of EMA in primary biliary cirrhosis, autoimmune hepatitis and primary sclerosing cholangitis did not contribute to CD diagnosis in a significant number of patients. 4) CD was most frequently diagnosed in the following liver diseases: cryptogenic hypertransaminasemia, autoimmune hepatitis, nodular regenerative hyperplasia and chronic hepatitis C and negative antimitochondrial primary biliary cirrhosis. 5) The removal of gluten from the diet contributed effectively to bring the hepatic enzymes levels back to normal in cases of cryptogenic hypertransaminasemia. However, the role of diet was difficult to evaluate in nodular regenerative hyperplasia, autoimmune hepatitis, alcohol disease and primary biliary cirrhosis groups due to the nature of the specific treatments and the natural course of the hepatic conditions.

Orientador: Maria Jaqueline Mamprim / Coorientador: Carlos Roberto Teixeira / Banca: Raquel Sartor Marcelino / Banca: Alessandra Melchert / Resumo: O atendimento de cães com sobrepeso e obesos na rotina clínica de animais de companhia tem sido cada vez mais frequente. O objetivo do presente estudo foi avaliar as alterações hepáticas parenquimatosas e hemodinâmicas, por meio da ultrassonografia bidimensional e Doppler, em três grupos de cães com diferentes escores de condição corporal - G1 (peso ideal/controle), G2 (sobrepeso) e G3 (obesos) -, sem doenças hepáticas por outras causas de base. Houve correlação positiva entre o G3 e os seguintes fatores: aumento da ecogenicidade do parênquima hepático, aumento das dimensões hepáticas, hiperlipidemia, e morfologia anormal da onda da veia hepática (VH). Diferença significativa na velocidade média do fluxo portal (Vméd.VP) foi observada entre G1 (17,23 ± 3,7cm/s) e G3 (13,66 ± 2,56cm/s), no volume do fluxo portal (VFVP) entre G1 (32,69 ± 11,86mL/min/kg) e G2 (19,58 ± 7,68mL/min/kg) e entre G1 e G3 (15,22 ± 8mL/min/kg), no índice hepatorrenal entre G2 (1,1 ± 0,29) e G3 (1,28 ± 0,22), nos níveis séricos de fosfatase alcalina (FA) entre G1 e G3, de gama glutamiltransferase (GGT) entre G1 e G2 e de colesterol entre G1 e os demais grupos. Não houve diferença significativa no índice de congestão portal (ICP), no índice de resistividade da artéria hepática (IRAH), no diâmetro e área da veia porta e nos níveis séricos de alanina aminotransferase (ALT) e triglicérides, entre os grupos. Concluiu-se que a obesidade pode levar ao desenvolvimento de alterações parenquimatosas e hemodinâmicas no fígado de cães / Abstract: Veterinary care of overweight and obese dogs in small animal clinical sciences has become increasingly common. The aim of the present study was to investigate, using two dimensional and Doppler sonography, parenchymal and hemodynamic changes in three groups of dogs, according to body score condition (G1, ideal body weight/control; G2, overweight; G3, obese), without hepatic diseases due to other causes. A positive correlation was found between G3 and the characteristics described below: increased liver echogenicity, hepatomegaly, hiperlipidemia and abnormal hepatic vein Doppler waveform pattern. There was statistical difference in the mean portal blood flow velocity between G1 (17,23 ± 3,7cm/s) and G3 (13,66 ± 2,56cm/s), in the portal blood flow between G1 (32,69 ± 11,86mL/min/kg) and G2 (19,58 ± 7,68mL/min/kg) and between G1 and G3 (15,22 ± 8mL/min/kg), in the hepatorenal index between G2 (1,1 ± 0,29) and G3 (1,28 ± 0,22), in serum levels of alkaline phosphatase (ALP) between G1 and G3, in levels of gamma glutamyltranspeptidase (GGT) between G1 and G2 and in levels of cholesterol between G1 and the other groups. There was no statistical difference in the portal vein congestion index, hepatic artery resistance index, portal vein diameter and area and on serum levels of alanine aminotransferase (ALT) and triglycerides between the groups. It was concluded that obesity may be responsible for the development of parenchymal and hemodynamic changes in dog's liver / Mestre

Le but de cette thèse était d’apporter un certain nombre d’améliorations techniques et méthodologiques à l’élastographie par résonance magnétique afin d’assurer le confort du patient, mais aussi de mieux comprendre les phénomènes impliqués dans le changement des propriétés mécaniques du foie. Ainsi, cette thèse a été construite de manière à améliorer les méthodes et les valider sur modèles animaux. La faisabilité clinique était étudiée en parallèle. Cette thèse repose principalement sur deux études expérimentales. En effet, différents modèles animaux ont été utilisés afin de mettre en relief le rôle de l’élastographie par résonance magnétique dans le diagnostic des maladies chroniques du foie. Ce doctorat a débouché sur deux publications en premier auteur. La première valide la méthode sur foie entier chez des rats avec fibrose hépatique induite au tétrachlorure de carbone et montre que les paramètres visco-élastiques sont corrélés à la quantité de fibrose. La deuxième étude, plus complexe, intègre différents modèles animaux afin d’apporter un début de réponse au rôle de l’élastographie par IRM dans le diagnostic des maladies à foie stéatosique. Il s’agissait de rats nourris avec une diète standard, une diète déficiente en choline jusqu’à 8 semaines pour induire une stéatohépatite, une diète enrichie en acide orotique pendant 2 semaines pour induire une stéatose simple, ou de rats ayant reçu une injection unique de CCl4 pour provoquer une lésion hépatique aigue. Les résultats principaux montraient une augmentation de la viscosité chez les rats à stéatose simple et une augmentation à la fois de l’élasticité et de la viscosité chez les rats avec lésion hépatique aigue. De plus, il a été montré chez les rats avec stéatohépatite une augmentation de l’élasticité et de la viscosité en l’absence de fibrose établie, alors que l’activation de la fibrogenèse était enclenchée, accompagnée d’une légère inflammation. L’analyse multivariée de tous les rats a montré que les changements d’élasticité sont principalement expliqués par l’activation des cellules stellaires (et donc de la fibrogenèse), alors que les changements de viscosité pourraient être expliqués par la présence de graisse. Ainsi, cette thèse a permis d’améliorer le confort du patient (matelas d’IRM et séquence rapide) pour faciliter l’intégration de cet examen en routine clinique, mais également de valider l’élastographie comme marqueur de la fibrose et des propriétés mécaniques des tissus. Pour finir, cette thèse a mis en avant le rôle potentiel de l’élastographie par IRM pour la détection précoce des patients à risque dans le cadre des stéatohépatites.

Graduation date: 1993

Chronic liver disease (CLD) is a leading cause of morbidity and mortality in HIV-infected individuals. The purposes of this study were to determine the prevalence and etiologies of CLD in an urban HIV-infected population and to identify CLD risk factors. We conducted a retrospective chart review of 799 HIV-infected patients seen at four New Haven health centers from 2002 to 2003. We applied the New Haven County Liver Study definition to identify patients with CLD. 65% were male, 44% were African American, and 23% were of Hispanic ethnicity. The mean age was 45 years. 30% had a history of alcohol abuse. 35% reported injection drug use as their HIV risk factor. Heterosexual contact and men having sex with men (MSM) were reported in 31% and 16% of cases. 50% of patients had a diagnosis of AIDS. 60% percent of patients had CLD. Over 50% of cases of CLD were attributed to chronic hepatitis C (HCV), either alone or with coexisting alcoholic liver disease. Alcoholic liver disease alone, hepatitis B virus (HBV), HAART-induced liver disease, and non-alcoholic liver disease (NAFLD) accounted for smaller percentages. 84% of patients were on HAART, but only 3.6% of patients with positive HCV or HBV serologies were on treatment for CLD. 75% of patients received pneumococcal and influenza vaccines, but only half of eligible patients received hepatitis A and B vaccines. In multivariate analysis, alcohol abuse and positive HCV status were associated with CLD. CLD is prevalent in our population. Preventive care and treatment for CLD are being overlooked in many. Vaccines, treatment for viral hepatitis, and strategies for reducing drug and alcohol abuse are priorities.

Thesis (Ph. D.)--University of Washington, 2002. / Vita. Includes bibliographical references (leaves 117-125).

published_or_final_version / abstract / Medicine / Doctoral / Doctor of Philosophy

Several trace metals have been shown to modify cell injury as indicated by reduction of observable pathological tissue changes after metal ion supplementation. An example of this is zinc ion induced protection against some of the liver injury caused by a single injection of carbon tetrachloride (CCl₄) to male Sprague-Dawley rats. Carbon tetrachloride liver injury is associated with membrane labilization as indicated by lysosomal and endoplasmic reticulum anomalies. Depressed hepatic levels of NADPH are observed during CCl₄ poisoning. Lipid metabolism is also impaired due to CCl₄ administration to animals. These biochemical manifestations of CCl₄ hepatotoxicity were studied in the presence of zinc ions in order to understand the mechanisms of the zinc protective effect against CCl₄ injury. The effect of zinc ions on the stability of rat liver lysomes was studied. Zinc was added by several methods: (1) feeding the animals a high zinc diet, (2) infusion of zinc into the liver in situ through the portal vein, or (3) by adding zinc to the lysosomal fraction either before or after isolation of this fraction from rat liver homogenates. By all techniques, addition of zinc reduced the release of β-glucuronidase from liver lysosomes, indicating increased stability of the suborganelles. The zinc induced protection against CCl₄ liver damage was evident in observations made using both light microscopy and electron microscopy. The increased release of lysosomal β-glucuronidase observed in the CCl₄ treated rats was significantly reduced by zinc administration. Also, decreases in microsomal protein synthesis and seromucoid levels due to the CCl₄ treatment were significantly ameliorated by zinc. Thus, disruption of lysosomal and endoplasmic reticulum membranes, one of the earliest signs of CCl₄ hepatotoxicity, appeared to be inhibited by pretreating the animals with zinc chloride. Depressed hepatic levels of NADPH are observed in rats administered CCl₄. Zinc chloride pretreatment of these rats significantly increased the NADPH levels in the liver. Since zinc ions bind NADPH, then the protective effect of zinc against CCl₄ toxicity may be due to stabilization of the pyridine nucleotide by zinc and the subsequent prevention of CCl₄ induced alterations of biochemical reactions dependent upon NADPH. Zinc chloride pretreatment of CCl₄ treated rats significantly reduced the CCl₄ induced hepatic triacylglycerol accumulation. Concomitant with this event is the appearance of elevated levels of newly synthesized triacylglycerols in the serum of the CCl₄ treated rats given zinc above that of the CCl₄ treated rats. Hepatic triacylglycerol synthesis is unchanged by CCl₄ or zinc treatment. Hepatic phospholipid levels which are depressed by the CCl₄ hepatotoxin are not affected by zinc treatment. However, the synthesis of phospholipids in the livers of rats treated with CCl₄ plus zinc is significantly increased. The lipid changes induced by CCl₄ and zinc dosing of rats are indicative of alterations in liver membranes thus affecting hepatic liver transport mechanisms. On the basis of the data presented, the effects of zinc ions on CCl₄ hepatotoxicity are discussed and applied to understanding the regulating role of metal ions in tissue injury. The protective effects of zinc ions against CCl₄ hepatotoxicity suggest a relationship between the zinc nutritional status of animals exposed to environmental contaminants, and the expression of the ensuing tissue damage.

The liver is a vital organ, playing numerous critical roles in the body. The liver's ability to perform essential functions is disturbed by injuries that are often associated with many complications such as calcification. Although many reports in the literature document observations of liver calcification, the mechanisms regulating this phenomenon remain unclear. Herein, we aim to investigate the cellular and molecular events that occur during pathological calcification of the liver. / To study the mechanisms of calcification, assessments included histological-staining, immunolabeling, and biochemical and electron microscopy analyses. The findings suggest that calcification may result from hydroxyapatite precipitation in necrotic or apoptotic hepatocytes. Similarly, calcification may be associated with differentiated myofibroblasts expressing bone matrix proteins downstream of TGFbeta signalling. / To identify specific protein regulators linked to the various stages in calcification, and to assess the protein composition of the tissue, a proteomic analysis was used. This analysis identified IQGAP1, an effector of the Rho-GTPases and a master regulator of cell adhesion and migration. IQGAP1 is strongly expressed in myofibroblasts, suggesting that IQGAP1 may be implicated in myofibroblasts migrating towards calcification. Studies on IQGAP1 interactions with its binding partners reveal that it is part of a protein complex that includes beta-catenin, an adhesion protein, and Rac1, a cytoskeletal regulator. These results suggest that IQGAP1 may play an important role in myofibroblast migration upon liver injury. / Having identified that activin and TGFbeta signalling are activated in myofibroblasts, we hypothesised that they may stimulate myofibroblast differentiation and proliferation. Studies using a C3H/10T1/2 cell model reveal that both activin and TGFbeta stimulate differentiation, but only activin induces cell proliferation in a Smad-independent fashion, which requires activation of the ERK/MAPK pathway. / In summary, this work provides new mechanistic insights on the global regulation of liver calcification. The various phases of this work collectively cover the central role of myofibroblasts in liver injury: association with calcification, rapid proliferation, differentiation to an activated form, and migration toward the injured area. The findings allow us to better understand the mechanisms by which liver myofibroblasts are regulated in a specific pathological context.