La muqueuse intestinale puis le foie sont en contact récurrent avec la flore microbienne issue du tube digestif. L’activation des récepteurs reconnaissant des motifs moléculaires microbiens (PRR :Pattern Recognition Receptors) constitue l’élément initial de la réponse inflammatoire de l’immunité innée et oriente le type d’activation de l’immunité adaptative. La première étape entraînera l’expression de médiateurs inflammatoire (cytokines (IL-1, IL-6, TNFα), activation de la réponse de phase aiguë) et le recrutement des cellules effectrices de l’immunité innée (neutrophiles, puis monocytes). L’équilibre entre la tolérance microbienne et l’exacerbation inflammatoire afin d’éliminer l’agent microbien et menant à la destruction tissulaire permet d’atteindre la situation d’homéostasie.<p>Si l’inflammation se perpétue, par exemple en cas de défaut de clairance de l’agent microbien ou de trouble de l’intégrité de la barrière muqueuse, l’inflammation peut devenir chronique. L’IL-6 exerce alors un rôle central dans la transition de l’immunité innée à l’immunité adaptative, en modulant différentiellement l’expression de chimiokines et l’apoptose des cellules immunes menant au remplacement de l’infiltrat neutrophilique par un infiltrat lympho-monocytaire. Par ailleurs, ce climat cytokinique particulier est propice au développement de lignées spécifiques de lymphocytes tels que les lymphocytes T CD4+ sécrétant de l’IL-17 (Th17). Ceux-ci, surtout étudiés dans les défenses anti-bactériennes et fungiques, et dans les maladies autoimmunes, ont été incriminés dans les phénomènes de cytotoxicité et de renouvellement inflammatoire par l’induction d’expression de chimiokines.<p><p>Une fois la barrière intestinale franchie, le foie est le premier organe en contact avec la flore microbienne issue de l’intestin. Certains TLRs ont été démontrés impliqués dans la physiopathologie de la stéatohépatite et dans le processus de fibrose. Ce climat constant d’exposition antigénique est associé en cas de maladie chronique du foie à une exacerbation d’expression de médiateurs infammatoires (IL-1, IL-6, TNFα).<p>Nous avons étudié la modulation d’expression des différents TLRs au cours d’un modèle de maladie alcoolique du foie chez la souris. Cette étude démontrait qu’il existait une majoration d’expression des TLR1, 2, 4, 6, 7, 8 et 9 dépendante du stress oxydatif suite à l’exposition chronique du foie à l’alcool ;celle-ci entraînant davantage de lésions hépatiques lors de l’injection des ligands respectifs de ces différents TLRs.<p>Dans ce contexte, nous avons également étudié l’expression des différentes sous-unités du récepteur à l’IL-6 au cours de deux maladies chroniques du foie chez l’homme :les maladies alcooliques du foie et l’hépatite C chronique. Nous avons mis en évidence que les taux plasmatiques d’IL-6 et de la forme soluble de gp130 augmentaient au cours des maladies chroniques du foie, de manière corrélée à la sévérité. Nous avons également démontré l’effet inhibiteur de sgp130 sur la réponse de phase aiguë dépendante du trans-signaling de l’IL-6 in vitro. Ces données suggèrent que sgp130 contribue au déficit de réponse de phase aiguë observé chez les patients atteints de cirrhose. <p><p>Par ailleurs, vu le contexte « cytokinique » chronique des maladies alcooliques du foie (IL-1 et IL-6), nous avons étudié l’activation de la voie de l’interleukine-17 et des Th17 au cours des maladies alcooliques du foie chez l’homme. Nous avons mis en évidence qu’il existait une activation de cellules circulantes sécrétant de l’IL-17 (comprenant des Th17) au cours de la cirrhose alcoolique stable. Par contre, au sein du foie, l’activation de cellules sécrétant de l’IL-17 était davantage augmentée lors de l’hépatite alcoolique. Nous avons également mis en évidence que les cellules stellées (cellules responsables de la fibrose hépatique) stimulées à l’IL-17 recrutaient les neutrophiles suite à l’expression d’IL-8 et de GROα. Cette nouvelle voie inflammatoire démontrée lors d’une maladie du foie chez l’homme met en évidence l’activation de la voie de l’IL-17 au cours des maladies alcooliques du foie et sa contribution potentielle au recrutement hépatique de neutrophiles au cours de l’hépatite alcoolique aiguë. Cette voie sera explorée dans l’avenir en termes de fonctionnalité et de potentielle cible thérapeutique. <p> <p>En conclusion, tout comme le suggère la littérature pour les maladies autoimmunes (maladie de Crohn, arthrite, sclérose en plaque), il semble que les maladies alcooliques du foie partagent avec ces dernières diverses caractéristiques inflammatoires. La flore microbienne intestinale participe à la physiopathologie des lésions hépatiques, de même que l’activation de PRR (TLR). Par ailleurs, un climat inflammatoire chronique (IL-6,…), contre-régulé par certains mécanismes (sgp130), est associé à la présence périphérique et hépatique de lymphocytes Th17. Cette dernière découverte ouvre de nouvelles perspectives dans la compréhension de la physiopathologie des maladies alcooliques du foie, et peut-être de nouvelles cibles thérapeutiques concernant l’hépatite alcoolique aiguë.<p> / Doctorat en Sciences médicales / info:eu-repo/semantics/nonPublished

Chronic alcohol use results in accelerated liver injury, leading to alcoholic steatohepatitis, cirrhosis, and hepatocellular carcinoma. However, due to the complex nature of this disease process, a central, druggable mechanism has remained elusive. microRNAs are potent post-transcriptional regulators of gene expression. A single miRNA has the ability to regulate hundreds of pathways simultaneously, defining cellular fate and function. microRNA-122 (miR-122), the most abundant miRNA in hepatocytes, has a demonstrated role as an tumor suppressor, regulator of hepatocyte metabolism, and hepatic differentiation. In this dissertation I demonstrate the role of miR-122 on alcoholic liver disease (ALD) pathogenesis over four parts. In chapter II, I will demonstrate chronic alcoholic patients, free of neoplastic changes, have a reduction of miR-122 and that this miRNA regulates HIF-1α, a determinant of ALD pathogenesis. In chapter III, using hepatocytetropic adeno-associated virus 8 (AAV8) vector, I demonstrate that miR-122 inhibition mimics ALD pathogenesis, and furthermore, using hepatocyte-specific HIF-1α-null (HIF1hepKO) mice that this phenomenon is HIF-1α dependent. Given this finding, in chapter IV, I demonstrate that ectopic expression of miR-122 in vivo can reverse alcoholinduced liver damage, steatosis, and inflammation by directly targeting HIF-1α. Finally, in chapter V, I present evidence that alcohol-induced dysregulation of grainyhead-like proteins 1 and 2 (GRHL2), mediate the inhibition of miR-122 at the transcriptional level. These findings dissect a novel mechanistic regulatory axis of miR-122 and indicate a potential opportunity for restoration of miR-122 as a therapy in early ALD.

published_or_final_version / abstract / Chinese Medicine / Master / Master of Philosophy

Li, Yanhong. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 192-212). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Wong Chun-kwan. / Thesis submitted in: December 1999. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2000. / Includes bibliographical references (leaves 127-137). / Abstracts in English and Chinese. / Abstract --- p.i / Acknowledgments --- p.viii / Tables of Contents --- p.ix / List of Figures --- p.xv / List of Tables --- p.xxvi / Chapter Chapter 1: --- INTRODUCTION --- p.1 / Chapter Chapter 2: --- LITERATURE REVIEW --- p.8 / Chapter 2.1 --- Toxicology --- p.8 / Chapter 2.1.1 --- Acute toxicity test --- p.8 / Chapter 2.1.2 --- Biochemical Analysis --- p.9 / Chapter 2.1.3 --- Organ weights --- p.10 / Chapter 2.2 --- Histology --- p.11 / Chapter 2.2.1 --- Light Microscope --- p.11 / Chapter 2.2.2 --- Electron Microscopy --- p.11 / Chapter 2.3 --- Tissue injury --- p.12 / Chapter 2.3.1 --- Free-radical mechanisms --- p.12 / Chapter 2.3.2 --- Lipid peroxidation --- p.13 / Chapter 2.4 --- Carbon tetrachloride (CC14) --- p.14 / Chapter 2.4.1 --- Mechanisms of carbon tetrachloride toxicity --- p.15 / Chapter 2.5 --- Trichloroethylene (TCE) --- p.18 / Chapter 2.5.1 --- Mechanisms of trichloroethylene toxicity --- p.21 / Chapter 2.6 --- Dimethyl sulfoxide (DMSO) --- p.25 / Chapter 2.7 --- N-acetylcysteine (NAC) --- p.27 / Chapter Chapter 3: --- MATERIALS AND METHODS --- p.28 / Chapter 3.1 --- Materials --- p.28 / Chapter 3.2 --- Methods --- p.31 / Chapter 3.2.1 --- Acute hepatotoxicity test on aqueous seaweed extracts --- p.31 / Chapter 3.2.1.1 --- Preparation of aqueous extracts of seaweed --- p.31 / Chapter 3.2.1.2 --- Experimental protocol --- p.31 / Chapter 3.2.1.3 --- Biochemical assays --- p.32 / Chapter 3.2.1.4 --- Organ weights --- p.36 / Chapter 3.2.1.5 --- Histopathological examination --- p.36 / Chapter 3.2.1.6 --- Statistical analysis --- p.36 / Chapter 3.2.2 --- Curative and preventive tests of seaweed aqueous extracts against the CCl4-induced hepatotoxicity --- p.37 / Chapter 3.2.2.1 --- Preparation of aqueous extracts of seaweed --- p.37 / Chapter 3.2.2.2 --- Experimental protocol --- p.37 / Chapter 3.2.2.3 --- Biochemical assays --- p.39 / Chapter 3.2.2.4 --- Organ weights --- p.39 / Chapter 3.2.2.5 --- Histopathological examination --- p.40 / Chapter 3.2.2.6 --- Statistical analysis --- p.41 / Chapter 3.2.3 --- Acute hepatotoxicity test of TCE in rats by oral and intraperitoneal routes --- p.42 / Chapter 3.2.3.1 --- Experimental protocol --- p.42 / Chapter 3.2.3.2 --- Biochemical assays --- p.43 / Chapter 3.2.3.3 --- Organ weights --- p.43 / Chapter 3.2.3.4 --- Histopathological examination --- p.44 / Chapter 3.2.3.5 --- Statistical analysis --- p.44 / Chapter 3.2.4 --- Curative and preventive tests of seaweed aqueous extracts against the TCE effective dose-induced toxicity --- p.44 / Chapter 3.2.4.1 --- Preparation of aqueous extracts of seaweed --- p.44 / Chapter 3.2.4.2 --- Experimental protocol --- p.45 / Chapter 3.2.4.3 --- Biochemical assays --- p.46 / Chapter 3.2.4.4 --- Organ weights --- p.46 / Chapter 3.2.4.5 --- Histopathological examination --- p.46 / Chapter 3.2.5 --- Antidotal effects of dimethyl sulfoxide (DMSO) and N-acetylcysteine (NAC) against CC14- and TCE- induced poisoning in rats --- p.47 / Chapter 3.2.5.1 --- Experimental protocol --- p.47 / Chapter 3.2.5.2 --- Biochemical assays --- p.48 / Chapter 3.2.5.3 --- Organ weights --- p.48 / Chapter 3.2.5.4 --- Histopathological examination --- p.49 / Chapter 3.2.6 --- Hepatoprotective effect of seaweeds' methanol extract against CC14- and TCE-induced poisoning in rats --- p.49 / Chapter 3.2.6.1 --- Preparation of methanol extracts of seaweed --- p.49 / Chapter 3.2.6.2 --- Experimental protocol --- p.50 / Chapter 3.2.6.3 --- Biochemical assays --- p.52 / Chapter 3.2.6.4 --- Organ weights --- p.52 / Chapter 3.2.6.5 --- Histopathological examination --- p.53 / Chapter Chapter 4 --- RESULTS --- p.54 / Chapter 4.1 --- Acute hepatotoxicity test on aqueous seaweed extracts --- p.54 / Chapter 4.1.1 --- The biochemical assays of the serum transaminase activity --- p.54 / Chapter 4.1.2 --- The organ weight (Aqueous seaweed crude extracts) --- p.56 / Chapter 4.2 --- Curative and preventive tests of seaweed aqueous extracts against the CCl4-induced hepatotoxicity --- p.58 / Chapter 4.2.1 --- The biochemical assays of the serum transaminase activity (Curative) --- p.58 / Chapter 4.2.2 --- The organ weight (Curative) --- p.60 / Chapter 4.2.3 --- The biochemical assays of the serum transaminase activity (Preventive) --- p.62 / Chapter 4.2.4 --- The organ weight (Preventive) --- p.64 / Chapter 4.3 --- Acute hepatotoxicity test of TCE in rats by oral and intraperitoneal routes --- p.66 / Chapter 4.3.1 --- Oral route --- p.66 / Chapter 4.3.1.1 --- One-time oral route --- p.66 / Chapter 4.3.1.2 --- Two-time oral route --- p.66 / Chapter 4.3.2 --- Intraperitoneal route --- p.66 / Chapter 4.3.3 --- Time course of the effective dose of 20% TCE in i.p. route --- p.67 / Chapter 4.4 --- Curative and preventive tests of seaweed aqueous extracts against the TCE effective dose-induced toxicity --- p.12 / Chapter 4.4.1 --- The biochemical assays of the serum transaminase activity (Curative) --- p.72 / Chapter 4.4.2 --- The organ weight (Curative) --- p.74 / Chapter 4.4.3 --- The biochemical assays of the serum transaminase activity (Preventive) --- p.76 / Chapter 4.4.4 --- The organ weight (Preventive) --- p.78 / Chapter 4.5 --- Antidotal effects of dimethyl sulfoxide (DMSO) and N-acetylcysteine (NAC) against CC14- and TCE-induced poisoning in rats --- p.80 / Chapter 4.5.1 --- The biochemical assays of the serum transaminase activity (Curative) --- p.80 / Chapter 4.5.2 --- The organ weight (Curative) --- p.82 / Chapter 4.5.3 --- The biochemical assays of the serum transaminase activity (Preventive) --- p.84 / Chapter 4.5.4 --- The organ weight (Preventive) --- p.86 / Chapter 4.6 --- Hepatoprotective effect of methanol extract of seaweed against CC14- and TCE-induced poisoning in rats --- p.88 / Chapter 4.6.1 --- The biochemical assays of the serum transaminase activity (Curative) --- p.88 / Chapter 4.6.2 --- The organ weight (Curative) --- p.89 / Chapter 4.7 --- Histopathological examinations --- p.90 / Chapter 4.7.1 --- Acute hepatotoxicity test on aqueous seaweed extracts --- p.91 / Chapter 4.7.2 --- Curative and preventive tests of seaweed aqueous extracts against the CC14-induced hepatotoxicity --- p.92 / Chapter 4.7.3 --- Acute hepatotoxicity test of TCE in rats by oral and intraperitoneal routes --- p.99 / Chapter 4.7.4 --- Curative and preventive tests of seaweed aqueous extracts against the TCE effective dose-induced toxicity --- p.100 / Chapter 4.7.5 --- Antidotal effects of dimethyl sulfoxide (DMSO) and N-acetylcysteine (NAC) against CC14- and TCE-induced poisoning in rats --- p.100 / Chapter 4.7.6 --- Hepatoprotective effect of methanol extract of seaweed against CC14- and TCE-induced poisoning in rats --- p.102 / Chapter Chapter 5 --- DISCUSSION --- p.106 / Chapter Chapter 6 --- CONCLUSION --- p.124 / REFERENCES --- p.127 / APPENDIX --- p.138

Shen, Jiayun. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 166-199). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

INTRODUÇÃO: A Doença Hepática Gordurosa (DHG) é considerada um dos grandes problemas de saúde pública do novo milênio, impulsionada sobretudo pela epidemia de obesidade. Está relacionada com alta morbidade, piora da qualidade de vida e custos econômicos para a sociedade. Existem duas formas histopatológicas iniciais inseridas dentro da DHG: a esteatose e a esteato-hepatite que podem evoluir para cirrose e hepatocarcinoma. A maior parte da literatura sobre epidemiologia da doença hepática gordurosa utiliza desenhos retrospectivos e meios diagnósticos não invasivos, embora o padrão ouro seja a biópsia hepática. Medidas preventivas e de controle adequados deverão ser tomadas baseadas em informações epidemiológicas envolvendo população \"sadia\". A excelente correspondência das alterações histopatológicas encontradas no cadáver em relação ao vivo permite extrapolar dados da frequência de esteatose e esteato-hepatite encontradas em uma população jovem vítima de morte violenta para uma população \"normal\". Contudo, não há estudo prévio que avalie frequência e fatores preditivos de esteatose e esteatohepatite em fígados de cadáver. O objetivo deste estudo foi avaliar a esteatose e a esteato-hepatite em fígados de adultos vítimas de morte violenta. MÉTODOS: As amostras foram coletadas de 224 adultos submetidos a autópsia forense a partir de setembro de 2011 a abril de 2013. Dados antropométricos e do fígado foram registrados. O exame histopatológico foi realizado em seis amostras obtidas com base em diferentes lóbulos de cada fígado. Cada amostra foi tratada com quatro corantes: hematoxilina-eosina, Perls, pricosirius e tricrômio de Masson. Desfechos de interesse foram a presença de esteatose, esteato-hepatite, fibrose e siderose. RESULTADOS: A amostra apresentou uma média de idade de 40 anos. A esteatose foi diagnosticada em 48,2% dos casos e esteato-hepatite em 2,7%. Uma alta prevalência de fígado gorduroso foi verificada entre homens e indivíduos mais velhos, sendo a faixa etária mais atingida entre 41-60 anos. Os fatores significativamente associados com o aumento do risco de esteatose foram circunferência abdominal (p < 0,001), IMC(p < 0,001), peso do fígado (p=0,002), assim como a presença de siderose (p=0.018). CONCLUSÃO: A alta prevalência de esteatose hepática foi detectada em biópsias pós-morte de uma população jovem. Uma vez que esta doença pode ter consequências clínicas severas, esse dado é importante para avaliar medidas preventivas para a doença hepática gordurosa e suas graves consequências / BACKGROUND: Fatty liver disease (FLD) is considered one of the major public health problems of the new millennium, driven mainly by the obesity epidemic. It is related to high morbidity, worsening of quality of life and economic costs to society. There are two initial histopathological forms inserted into the FLD: steatosis and steatohepatitis which can progress to cirrhosis and hepatocellular carcinoma. Most of the literature on epidemiology of FLD using retrospective drawings and non invasive means, but the gold standard is a liver biopsy. Preventive measures and adequate control should be taken based on epidemiological information involving population \"healthy\". The excellent correlation of histopathologic changes found in the body in live relationship allows extrapolating data from steatosis to steatohepatitis and often found in a population young victim of violent death to a \"normal\" population. However, no previous study to assess the frequency and predictors of steatosis and steatohepatitis in cadaver livers. The aim of this study is to evaluate steatosis and steatohepatitis in livers of adult victims of violent death. METHODS: Specimens were collected from 224 adults undergoing forensic autopsy from September 2011 to April 2013. Anthropometric and liver data were recorded. Histopathological examination was performed on six biopsies obtained from different lobes of each liver. Each sample was treated with 4 stains: hematoxylin-eosin, Perls, pricosirius and trichrome Masson. Outcomes of interest were the presence of steatosis, steatohepatitis, fibrosis and siderosis. RESULTS: The sample had an average age of 40 years. The steatosis was detected in 48.2% of cases and 2.7% steatohepatitis. A high prevalence of fatty liver was observed among men and older individuals, the age group most affected between 41-60 years. The factors significantly associated with increased risk of steatosis were waist circumference (p < 0.001), BMI (p < 0.001), liver weight (p = 0.002) as well as the presence of siderosis ( p = 0.018). CONCLUSION: The high prevalence of hepatic steatosis was detected in postmortem biopsy of a young population. Since this disease can have severe clinical consequences, this data is important to evaluate preventive measures for fatty liver disease and its serious consequences

Keung Hoi Yee. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 151-174). / Abstracts in English and Chinese. / Acknowledgement --- p.I / Abstract --- p.II / 摘要 --- p.V / Content --- p.VII / List of tables --- p.XIII / List of figures --- p.XIV / Abbreviations --- p.XVII / Chapter Chapter 1 --- General Introduction --- p.1 / Chapter 1.1 --- Biology of Pleurotus tuber-regiun (Ptr) --- p.1 / Chapter 1.1.1 --- Ptr grown in the wild --- p.1 / Chapter 1.1.2 --- Cultivation of Ptr --- p.2 / Chapter 1.2 --- Functional food and pharmaceutical application of Ptr sclerotium --- p.3 / Chapter 1.2.1 --- Traditional food and medicinal uses of Ptr sclerotium --- p.3 / Chapter 1.2.2 --- Nutritional value and chemical composition --- p.4 / Chapter 1.2.3 --- Anti-tumor activity --- p.7 / Chapter 1.2.4 --- Anti-viral activity --- p.8 / Chapter 1.2.5 --- Immunologic function --- p.8 / Chapter 1.2.6 --- Pharmaceutical application --- p.9 / Chapter Chapter 2 --- Toxicological evaluation on Ptr sclerotium --- p.11 / Chapter 2.1 --- Introduction --- p.11 / Chapter 2.1.1 --- Toxicological concern of Ptr sclerotium --- p.11 / Chapter 2.1.2 --- Toxicological study --- p.12 / Chapter 2.1.3 --- Biochemical methods for toxicological evaluation --- p.14 / Chapter 2.1.3.1 --- Serum enzyme activities --- p.15 / Chapter 2.1.3.2 --- Other serum analytes --- p.17 / Chapter 2.1.4 --- Histopathological study --- p.20 / Chapter 2.1.5 --- Acute toxicity --- p.21 / Chapter 2.1.6 --- Sub-acute and sub-chronic toxicity --- p.23 / Chapter 2.1.7 --- Objectives --- p.26 / Chapter 2.2 --- Materials and Methods --- p.27 / Chapter 2.2.1 --- Sample materials and chemicals --- p.27 / Chapter 2.2.2 --- Acute toxicity test --- p.27 / Chapter 2.2.2.1 --- Diet and animals --- p.27 / Chapter 2.2.2.2 --- Experimental design --- p.28 / Chapter 2.2.2.3 --- Calculation of sclerotium intake dose --- p.29 / Chapter 2.2.2.4 --- Biochemical assays --- p.30 / Chapter 2.2.2.5 --- Histopathological examination --- p.31 / Chapter 2.2.3 --- Sub-acute and sub-chronic toxicity tests --- p.32 / Chapter 2.2.3.1 --- Diet Preparation --- p.32 / Chapter 2.2.3.2 --- Experimental design --- p.32 / Chapter 2.2.3.3 --- Biochemical assays --- p.36 / Chapter 2.2.3.4 --- Organ weight --- p.40 / Chapter 2.2.3.5 --- Histopathological examination --- p.41 / Chapter 2.2.4 --- Statistical analyses --- p.41 / Chapter 2.3 --- Results and Discussion --- p.42 / Chapter 2.3.1 --- Acute toxicity test --- p.42 / Chapter 2.3.1.1 --- Food consumption --- p.43 / Chapter 2.3.1.2 --- Serum transaminase activities --- p.44 / Chapter 2.3.1.3 --- Histopathology --- p.45 / Chapter 2.3.1.4 --- NOAEL --- p.45 / Chapter 2.3.2 --- Sub-acute toxicity test --- p.50 / Chapter 2.3.2.1 --- Body weight gain --- p.50 / Chapter 2.3.2.2 --- Biochemical assays --- p.51 / Chapter 2.3.2.3 --- Organ per body weight and histopathology --- p.52 / Chapter 2.3.2.4 --- Effects of Ptr sclerotial diets --- p.53 / Chapter 2.3.3 --- Sub-chronic toxicity test --- p.59 / Chapter 2.3.3.1 --- Food and energy consumption --- p.59 / Chapter 2.3.3.2 --- Biochemical assays --- p.63 / Chapter 2.3.3.3 --- Organ per body weight --- p.67 / Chapter 2.3.3.4 --- Body weight increase --- p.75 / Chapter 2.3.3.5 --- NOAEL --- p.80 / Chapter 2.4 --- Summary --- p.81 / Chapter Chapter 3 --- Hepatoprotection of Ptr sclerotium --- p.82 / Chapter 3.1 --- Introduction --- p.82 / Chapter 3.1.1 --- Hepatotoxicity --- p.82 / Chapter 3.1.2 --- Potential hepatoprotection effect of Ptr sclerotium --- p.83 / Chapter 3.1.3 --- Toxicity of CC14 --- p.85 / Chapter 3.1.4 --- Toxicity of AFB! --- p.89 / Chapter 3.1.5 --- Bioactivity of chlorophyllin --- p.92 / Chapter 3.1.6 --- Comet assay --- p.93 / Chapter 3.1.7 --- Objectives --- p.98 / Chapter 3.2 --- Materials and Methods --- p.99 / Chapter 3.2.1 --- Sample materials and chemicals --- p.99 / Chapter 3.2.2 --- Curative and preventive tests of Ptr sclerotium against CCl4-induced hepatotoxicity --- p.99 / Chapter 3.2.2.1 --- Animal and diets --- p.99 / Chapter 3.2.2.2 --- Dose-response of CCl4 on rat model --- p.100 / Chapter 3.2.2.3 --- Biochemical assays --- p.100 / Chapter 3.2.2.4 --- Curative hepatoprotection test on Ptr --- p.101 / Chapter 3.2.2.5 --- Preventive hepatoprotection test on Ptr --- p.101 / Chapter 3.2.3 --- Preventive tests of Ptr sclerotium against AFB1-induced hepato- and geno-toxicity --- p.103 / Chapter 3.2.3.1 --- Dose-response of AFB1 on rat model --- p.103 / Chapter 3.2.3.2 --- Preventive test of Ptr against AFB1 --- p.103 / Chapter 3.2.3.3 --- Biochemical assays --- p.105 / Chapter 3.2.3.4 --- Histopathological examination --- p.105 / Chapter 3.2.4 --- Comet assay --- p.106 / Chapter 3.2.4.1 --- Reagent preparations --- p.106 / Chapter 3.2.4.2 --- Procedures --- p.107 / Chapter 3.2.5 --- Statistical analyses --- p.110 / Chapter 3.3 --- Results and Discussion --- p.111 / Chapter 3.3.1 --- Curative and preventive tests of Ptr sclerotium against CCl4-induced hepatotoxicity --- p.112 / Chapter 3.3.1.1 --- Dose-response of CCl4 on rat model --- p.112 / Chapter 3.3.1.2 --- Curative test of Ptr sclerotium against CCl4-induced hepatotoxicity --- p.116 / Chapter 3.3.1.3 --- Preventive test of Ptr sclerotium against CCl4-induced hepatotoxicity --- p.121 / Chapter 3.3.2 --- Preventive tests of Ptr sclerotium against AFB1-induced hepato- and geno-toxicity --- p.126 / Chapter 3.3.2.1 --- Dose-response of AFB1 on rat model --- p.126 / Chapter 3.3.2.2 --- Preventive test of Ptr sclerotium against AFB1-induced geno- and hepatotoxicity --- p.134 / Chapter 3.3.2.3 --- CHL versus 30% Ptr sclerotial diet --- p.137 / Chapter 3.3.3 --- A comparison of the hepatotoxicity of CC14 and AFB1 --- p.142 / Chapter 3.4 --- Summary --- p.147 / Chapter Chapter 4 --- Conclusions and future work --- p.148 / References --- p.151 / Related publication --- p.175

Submitted by Fabíola Silva (fabiola.silva@famerp.br) on 2017-08-03T12:25:59Z No. of bitstreams: 1 andrerodrigueirocpdeoliveira_tese.pdf: 1590220 bytes, checksum: c85526dbe8e00ebf97a87b9215d49485 (MD5) / Made available in DSpace on 2017-08-03T12:25:59Z (GMT). No. of bitstreams: 1 andrerodrigueirocpdeoliveira_tese.pdf: 1590220 bytes, checksum: c85526dbe8e00ebf97a87b9215d49485 (MD5) Previous issue date: 2015-11-27 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / Fundação de Amparo à Pesquisa do Estado de São Paulo - FAPESP / Introduction: Hepatocellular carcinoma (HCC) is a primary liver tumor and the sixth most common type of cancer. Liver cirrhosis (LC) and viral hepatitis are their main risk factors. Injured tissues and development process tissues need new blood vessels, i. e., angiogenic process, that initiates by means of vascular endothelial growth factors secretion, that is translated by VEGFA gene and regulated by non-coding RNA, miRNA, that consists of a single strand of approximately 22 nucleotides and has the function of binding to mRNA strand, to inhibit protein translation. Objectives: To quantify VEGFA gene expression and 17 miRNAs predicted as its regulator in HCC and liver cirrhosis; to assess VEGFA gene expression in relation to miRNAs expression and quantify VEGFA protein expression in HCC and liver cirrhosis tissues. Patients and methods: Sixteen samples of HCC tissue, 23 samples of liver cirrhosis tissue and 12 samples of normal liver tissue were collected. Total RNA and proteins were isolated by mirVana PARIS kit. VEGFA gene mRNA quantification and 17 miRNAs predicted as regulators expression was performed by real time quantitative PCR. Customized plates, TaqMan miRNA custom plate, were used for miRNAs. Statistical analysis was performed by one sample t test, Wilcoxon tests and Mann-Whitney test. Results: VEGFA gene expression was found downregulated in HCC and there is negative correlation between gene expression and tumor extension. MicroRNA hsa-miR-637 has showed significance and was down-regulated on samples (mean 0.2003, p=0.0004) in LC, whereas miRNAs hsa-miR-15b (median 0.04015; p=0.0005), hsa-miR-125b (median 0.01876; p=0.0005), hsa-miR-423 (median 0.02650; p=0.0005), hsa-miR-424 (median 0.00462; p=0.0156), hsa-miR-494 (median 0.00877; p=0.0010), hsa-miR-497 (median 0.04487; p=0.0005), hsa-miR-612 (median 0.00679; p=0.0039) and hsamiR-637 (median 0.00166; p=0.0039) are down-regulated in HCC. Conclusions: VEGFA gene expression is down-regulated in HCC. MicroRNA hsa-miR-637 is down-regulated in LC. MicroRNAs hsa-miR-15b, hsa-miR-125b, hsa-miR-423, hsa-miR-424, hsa-miR-494, hsa-miR-497, hsa-miR-612 e hsa-miR-637 are down-regulated in HCC. VEGFA protein expression is low in HCC, corroborating to gene expression found in HCC. / Introdução: O Carcinoma hepatocelular (CHC) é o tipo de tumor primário do fígado e o quinto tipo de câncer mais comum. A cirrose hepática (CH) e as hepatites virais são seus principais fatores de risco. Tecidos lesionados e em processo de crescimento necessitam de novos vasos sanguíneos, ou seja, processo angiogênico, que é iniciado por meio da secreção de fatores de crescimento endoteliais vasculares. Este é produzido a partir do gene VEGFA e regulado por RNA não codificante, o microRNA, que é constituído por uma fita simples de aproximadamente 22 nucleotídeos e tem a função de ligação ao mRNA, para inibição da tradução de proteínas. Objetivos: Quantificar a expressão do gene VEGFA e 17 miRNAs preditos como seus reguladores em CHC e cirrose hepática; avaliar a expressão do gene VEGFA em relação à expressão dos miRNAs e quantificar a expressão das proteínas VEGFA em CHC e cirrose hepática. Casuística e métodos: Foram coletadas 16 amostras de tecido de CHC, 23 amostras de tecido cirrose hepática e 12 amostras de tecido hepático normal para controle. A extração de RNA e proteínas totais foi realizada por meio do kit mirVana PARIS. A quantificação do mRNA do gene VEGFA e dos 17 miRNAs selecionados preditos como reguladores do gene foi realizada por meio da técnica de PCR quantitativa em tempo real. Para os miRNAs foram utilizadas placas customizadas TaqMan miRNA custom plates. A análise estatística foi realizada por teste t de uma amostra, teste de Wilcoxon e teste de Mann-Whitney. Resultados: O gene VEGFA foi encontrado em expressão diminuída em CHC e há correlação negativa entre a expressão e a extensão do tumor. O miRNA hsa-miR-637 mostrou significância e sua expressão foi encontrada diminuída nas amostras (média=0,2003, p=0,0004) em CH, enquanto que os miRNAs hsa-miR-15b (mediana 0,04015; p=0,0005), hsa-miR- 125b (mediana 0,01876; p=0,0005), hsa-miR-423 (mediana 0,02650; p=0,0005), hsa-miR-424 (mediana 0,00462; p=0,0156), hsa-miR-494 (mediana 0,00877; p=0,0010), hsa-miR-497 (mediana 0,04487; p=0,0005), hsa-miR-612 (mediana 0,00679; p=0,0039) e hsa-miR-637 (mediana 0,00166; p=0,0039) estão em expressão diminuída em CHC. Conclusão: A expressão do gene VEGFA encontra-se diminuída em CHC. O microRNA hsa-miR-637 encontra-se em expressão diminuída em CH. Os miRNAs hsa-miR-15b, hsa-miR-125b, hsa-miR- 423, hsa-miR-424, hsa-miR-494, hsa-miR-497, hsa-miR-612 e hsa-miR-637 encontram-se em expressão diminuída em CHC. A expressão das proteínas VEGFA está diminuída em carcinoma hepatocelular, corroborando com o encontrado para a expressão gênica em CHC.

Li Mi. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 125-136). / Abstracts in English and Chinese. / Abstract --- p.i / 論文摘要 --- p.iv / Publications --- p.vi / Acknowledgement --- p.vii / Abbreviations --- p.viii / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Pyrrolizidine alkaloids --- p.1 / Chapter 1.1.1 --- Distribution and plant sources --- p.1 / Chapter 1.1.2 --- Structures and nomenclature --- p.3 / Chapter 1.2 --- PA-containing Chinese medicinal herbs --- p.6 / Chapter 1.3 --- PA-induced toxicity。 --- p.7 / Chapter 1.3.1 --- Acute toxicity and chronic toxicity --- p.7 / Chapter 1.3.2 --- Genotoxicity --- p.8 / Chapter 1.3.3 --- Tumorigenicity --- p.8 / Chapter 1.3.4 --- Hepatotoxicity --- p.8 / Chapter 1.3.5 --- Mechanism of toxic effects --- p.9 / Chapter 1.3.5.1 --- Metabolic pathways --- p.10 / Chapter 1.3.5.2 --- Liver tissue-bound pyrroles --- p.16 / Chapter 1.3.5.3 --- Metabolizing enzymes --- p.17 / Chapter 1.3.5.3.1 --- Phase I metabolizing enzymes --- p.17 / Chapter 1.3.5.3.2 --- Phase II metabolizing enzymes --- p.18 / Chapter 1.3.5.4 --- Species and gender specificity toward toxicity --- p.19 / Chapter 1.3.5.5 --- Structure-activity relationships --- p.20 / Chapter 1.4 --- Prevention of PAs-induced toxicity --- p.23 / Chapter 1.4.1 --- Significance of prevention in humans --- p.23 / Chapter 1.4.2 --- Regulations toward preventing toxicity induced by PAs --- p.24 / Chapter 1.5 --- Aim of the present study --- p.25 / Chapter Chapter 2 --- Qualitative and Quantitative Analysis of PA-containing Chinese Medicinal Herbs --- p.26 / Chapter 2.1 --- Materials and equipments --- p.27 / Chapter 2.1.1 --- Herbal materials --- p.27 / Chapter 2.1.2 --- Chemicals and solvents --- p.27 / Chapter 2.1.3 --- Equipment and instrumentation --- p.27 / Chapter 2.2 --- Preparation of herbal extracts。 --- p.28 / Chapter 2.2.1 --- Crude herbal extract --- p.28 / Chapter 2.2.2 --- Total pyrrolizidine alkaloid extract --- p.28 / Chapter 2.3 --- Qualitative and quantitative analysis of Ligularia hodgsonii --- p.29 / Chapter 2.3.1 --- Methods --- p.29 / Chapter 2.3.1.1 --- HPLC-UV condition --- p.29 / Chapter 2.3.1.2 --- HPLC-MS condition --- p.29 / Chapter 2.3.1.3 --- Calibration curve for clivorine --- p.29 / Chapter 2.3.1.4 --- Recovery test --- p.30 / Chapter 2.3.1.5 --- Sample test --- p.30 / Chapter 2.3.2 --- Results and discussions --- p.30 / Chapter 2.3.2.1 --- Qualitative analysis of PAs in Ligularia hodgsonii --- p.30 / Chapter 2.3.2.2 --- Calibration curve for clivorine --- p.35 / Chapter 2.3.2.3 --- Result of the recovery test --- p.37 / Chapter 2.3.2.4 --- Quantification of PAs in Ligularia hodgsonii --- p.37 / Chapter 2.4 --- Qualitative and quantitative analysis of Tussilago farfara --- p.37 / Chapter 2.4.1 --- Methods --- p.38 / Chapter 2.4.1.1 --- HPLC-UV condition --- p.38 / Chapter 2.4.1.2 --- HPLC-MS condition --- p.39 / Chapter 2.4.1.3 --- Calibration curve for senkirkine --- p.39 / Chapter 2.4.1.4 --- Recovery test --- p.39 / Chapter 2.4.1.5 --- Sample test --- p.39 / Chapter 2.4.2 --- Results and discussions --- p.40 / Chapter 2.4.2.1 --- Qualitative analysis of senkirkine in Tussilago farfara --- p.40 / Chapter 2.4.2.2 --- Calibration curve for senkirkine --- p.42 / Chapter 2.4.2.3 --- Result of the recovery test --- p.42 / Chapter 2.4.2.4 --- Quantification of senkirkine in Tussilago farfara --- p.43 / Chapter 2.5 --- Qualitative and quantitative analysis of Gynura segetum --- p.43 / Chapter 2.5.1 --- Methods --- p.43 / Chapter 2.5.1.1 --- HPLC-UV condition --- p.43 / Chapter 2.5.1.2 --- HPLC-MS condition --- p.44 / Chapter 2.5.1.3 --- Calibration curves for senecionine and seneciphylline --- p.44 / Chapter 2.5.1.4 --- Recovery test… --- p.44 / Chapter 2.5.1.5 --- Sample test --- p.44 / Chapter 2.5.2 --- Results and discussions。 --- p.45 / Chapter 2.5.2.1 --- Qualitative analysis of PAs in Gynura segetum --- p.45 / Chapter 2.5.2.2 --- Calibration curves for senecionine and seneciphylline --- p.49 / Chapter 2.5.2.3 --- Result of the recovery test --- p.49 / Chapter 2.5.2.4 --- Quantification of PAs in Gynura segetum --- p.49 / Chapter 2.6 --- Qualitative and quantitative analysis of Crotalaria sessiliflora --- p.50 / Chapter 2.6.1 --- Methods --- p.50 / Chapter 2.6.1.1 --- HPLC-UV condition --- p.50 / Chapter 2.6.1.2 --- HPLC-MS condition --- p.50 / Chapter 2.6.1.3 --- Calibration curve --- p.51 / Chapter 2.6.1.4 --- Recovery test --- p.51 / Chapter 2.6.1.5 --- Sample test --- p.51 / Chapter 2.6.2 --- Results and discussions --- p.52 / Chapter 2.6.2.1 --- Qualitative analysis of monocrotaline in Crotalaria sessiliflora --- p.52 / Chapter 2.6.2.2 --- Calibration curve for monocrotaline --- p.54 / Chapter 2.6.2.3 --- Result of the recovery test --- p.54 / Chapter 2.6.2.4 --- Quantification of PAs in Crotalaria sessiliflora --- p.55 / Chapter 2.7 --- Qualitative and quantitative analysis of Senecio scandens --- p.55 / Chapter 2.7.1 --- Methods --- p.55 / Chapter 2.7.1.1 --- HPLC-UV condition --- p.55 / Chapter 2.7.1.2 --- HPLC-MS condition --- p.56 / Chapter 2.7.1.3 --- Sample test --- p.56 / Chapter 2.7.2 --- Results and discussions --- p.56 / Chapter 2.7.2.1 --- Qualitative analysis of PAs in Senecio scandens --- p.56 / Chapter 2.7.2.2 --- Quantification of PAs in Senecio scandens --- p.59 / Chapter Chapter 3 --- Hepatotoxicity Induced by PA-containing Chinese Medicinal Herbs --- p.60 / Chapter 3.1 --- Materials and methods --- p.62 / Chapter 3.1.1 --- Reagents --- p.62 / Chapter 3.1.2 --- Animal models --- p.62 / Chapter 3.1.3 --- Determination of the serum ALT activity --- p.64 / Chapter 3.1.4 --- Determination of hepatic GSH level --- p.68 / Chapter 3.1.5 --- Quantitation of liver tissue-bound pyrroles --- p.69 / Chapter 3.1.6 --- Histological assessment of liver morphological changes --- p.70 / Chapter 3.1.7 --- Assessment of hepatocytes apoptosis --- p.71 / Chapter 3.1.8 --- Statistical analysis --- p.72 / Chapter 3.2 --- Results and discussion --- p.72 / Chapter 3.2.1 --- Calibration curves --- p.72 / Chapter 3.2.1.1 --- Calibration curve for the determination of serum ALT activity --- p.72 / Chapter 3.2.1.2 --- Calibration curve of determination of hepatic GSH level --- p.73 / Chapter 3.2.2 --- Hepatotoxicity Study of Crotalaria sessiliflora --- p.74 / Chapter 3.2.2.1 --- Hepatotoxicity at 24 hrs after treatment --- p.74 / Chapter 3.2.2.1.1 --- Correlation between dosage of monocrotaline in Crotalaria sessiliflora and amount of liver tissue-bound pyrroles --- p.74 / Chapter 3.2.2.1.2 --- Effects of Crotalaria sessiliflora on the serum ALT activity --- p.79 / Chapter 3.2.2.1.3 --- The correlation between the elevated level of ALT activity and apoptosis of liver cells --- p.85 / Chapter 3.2.2.1.4 --- Effects of Crotalaria sessiliflora on the hepatic GSH level --- p.86 / Chapter 3.2.2.1.5 --- Histological changes of liver sections --- p.89 / Chapter 3.2.2.2 --- Hepatotoxicity within 4 days after administration --- p.92 / Chapter 3.2.2.3 --- Sub-acute hepatotoxicity within 14 days after administration --- p.93 / Chapter 3.2.2.4 --- Conclusion in hepatotoxicity study of Crotalaria sessiliflora --- p.99 / Chapter 3.2.3 --- Hepatotoxicity Study of Gynura segetum --- p.102 / Chapter 3.2.3.1 --- Correlation between the dosage of PAs present in Gynura segetum and the amount of liver tissue-bound pyrroles --- p.102 / Chapter 3.2.3.2 --- "Effects of Gynura segetum on serum ALT activity, hepatic GSH level and morphological changes of liver" --- p.104 / Chapter 3.2.4 --- Hepatotoxicity Study of Ligularia hodgsonii --- p.108 / Chapter 3.2.4.1 --- Correlation between the dosage of PAs present in Ligularia hodgsonii and the formation of liver tissue-bound pyrroles --- p.108 / Chapter 3.2.4.2 --- Effects of Ligularia hodgsonii on serum ALT activity and hepatic GSH level --- p.111 / Chapter 3.2.5 --- Hepatotoxicity Study of Tussilago farfara --- p.113 / Chapter 3.2.6 --- Hepatotoxicity Study of PA-containing medicinal herbs --- p.115 / Chapter 3.2.6.1 --- Correlation between formation of liver tissue-bound pyrroles and elevated serum ALT level --- p.115 / Chapter 3.2.6.2 --- Correlation between dosage of PAs and amount of liver tissue-bound pyrroles --- p.117 / Chapter 3.2.7 --- Test of Liver Tissue-bound Pyrroles as a biomarker using Senecionis scandentis --- p.118 / Chapter 3.2.8 --- Conclusions --- p.120 / Chapter Chapter 4 --- General Conclusions --- p.121 / Chapter 4.1 --- Qualitative and quantitative analysis of five PA-containing medicinal herbs --- p.121 / Chapter 4.2 --- Hepatotoxicity induced by PA-containing medicinal herbs in rats --- p.122 / Chapter 4.3 --- The correlation between hepatotoxicity induced by PA-containing medicinal herbs and the formation of liver tissue-bound pyrroles --- p.123 / Chapter 4.4 --- Threshold of the amount of liver tissue-bound pyrroles related to the hepatotoxicity induced by PA-containing medicinal herbs --- p.123 / References --- p.125