O aumento na incidência do carcinoma hepatocelular (CHC) tem sido atribuído ao aumento da obesidade, diabetes e doença hepática gordurosa não alcoólica. (DHGNA), estando ainda por ser melhor esclarecidos vários aspectos histopatológicos e imuno-histoquímicos. O objetivo deste estudo é avaliar aspectos clínicos e patológicos de pacientes com CHC secundário a DHGNA, assim como relacionar a marcadores imuno-histoquímicos de classe proliferativa. Avaliamos 35 espécimes de CHC de 21 pacientes diagnosticados com DHGNA submetidos a ressecção hepática (12 pacientes) ou a transplante hepático (8 pacientes) ou ambos (1 paciente), de 2005 a 2015. Dados demográficos, clínicos e bioquímicos foram relacionados a características histológicas e reatividade imuno-histoquímica para K19, marcando características de células progenitoras e Ki-67, marcando as células em ciclo celular. Um total de 35 nódulos foram detectados em 21 pacientes. A cirrose estava presente em 12 casos (7 F4A x 4F4B x 1F4C de acordo com estadiamento de Laennec) e 9 pacientes não apresentavam cirrose (estadiamento DHGNA: F2: 6pts, F3 = 3pts). A idade variou de 50 a 77 anos e 16 pacientes eram do sexo masculino (76%). Dezesseis pacientes (76%) apresentavam diabetes mellitus, 17 pacientes (81%) apresentavam hipertensão arterial e 19 pacientes (90%) tinham IMC superior a 25 kg / m2. O CHC ocorreu em 8 pacientes CHILD A, 4 CHILD B e em 9 pacientes sem cirrose. O nível de alfa-fetoproteína foi normal em 13 (62%) pacientes. Dentre os critérios histológicos, 25 (70%) nódulos foram diagnosticados como \"CHC esteatohepatítico\". Embora 63% tenham sido pouco diferenciados (G.3/G.4) de acordo com Edmondson & Steiner (1954), apenas 21% apresentaram níveis elevados de Ki-67 ( > 10%). No caso da K19, também 21% dos pacientes apresentaram expressão positiva ( > 5%), e foi associado a maior inflamação intratumoral (G 2/3). Curiosamente, 75% dos pacientes com alta expressão de Ki-67 ( > 10%) não eram cirróticos. Em conclusão: 1. Nesta casuística cirúrgica, o CHC relacionado com DHGNA foi encontrado em não cirróticos em 42% dos casos, com nível normal de alfafetoproteína em 62%. 2. Os marcadores histológicos de \"CHC esteatohepatítico\" estiveram altamente prevalentes. 3. A imunoexpressão positiva de K19 e Ki-67 ocorreu em apenas 21% dos pacientes, o que pode sugerir que o CHC na síndrome metabólica pode ser preferencialmente \"um subtipo inflamatório e não proliferativo de CHC\" / The increase incidence of hepatocellular carcinoma (HCC) has been attributed to the increase in obesity, diabetes and non-alcoholic fatty liver disease. (NAFLD), and several histopathological and immunohistochemical aspects are still to be better clarified. The aim of this study is to assess clinical and pathological aspects of patients with HCC secondary to NAFLD as well as to related to immunohistochemical markers of proliferative class. We evaluated 35 HCC specimens from 21 patients diagnosed with NAFLD undergoing liver resection (12 patients) or liver transplantation (8 patients) or both (1 patient) from 2005 to 2015. Demographic, clinical and biochemical data were related to histological features and immunohistochemical reactivity for K19, marking characteristics of progenitor cells and Ki-67, marking the cells in cell cycle. A total of 35 nodules were detected from 21 patients. Cirrhosis was present in 12 cases (7 F4A x 4F4B x 1F4C according to Laennec Staging) and 9 patients did not have cirrhosis (NAFLD staging: F2: 6pts, F3=3pts). Ages ranged from 50 to 77 years and 16 patients were male (76%). Sixteen patients (76%) had diabetes mellitus, 17 patients (81%) had arterial hypertension and 19 patients (90%) had BMI above 25kg/m2. HCC occurred in 8 patients Child A, 4 Child B and in 9 patients without cirrhosis. Alpha-fetoprotein level was normal in 13 (62%) patients. Among the histological criteria, 25 (70%) nodules were diagnosed as \"steatohepatitic HCC\". Although 63% were poorly differentiated (G.3/ G.4) according to Edmondson & Steiner (1954), only 21% presented high levels of Ki-67 ( > 10%). In the case of K19, 21% of patients presented positive expression (> 5%), and was associated with greater intratumoral inflammation (G 2/3). Interestingly, 75% of the patients with high Ki67 expression ( > 10%) were non-cirrhotic. In conclusion: 1. In this surgical series, HCC related to NAFLD was found in non-cirrhotic patients in 42% of cases, with a normal level of alpha-fetoprotein in 62%. 2. Histological markers of \"steatohepatitic HCC\" were highly prevalent. 3. Positive immunoexpression of K19 and Ki-67 occurred in only 21% of patients, which might suggest that HCC in metabolic syndrome might be preferentially \"an inflammatory, non-proliferative subtype of HCC\"

A doença hepática gordurosa não alcoólica (DHGNA) consiste na alteração morfofisiológica do fígado decorrente do acúmulo de lipídios nos hepatócitos. O desenvolvimento de DHGNA pode estar associado à obesidade e o diabetes tipo 2, e um dos possíveis mecanismos mediadores é a hiperatividade da alça ECA/Ang II/AT1 do sistema renina angiotensina (SRA). Por outro lado, evidências mais recentes mostraram que a ativação da alça ECA2/Ang 1-7/Mas do SRA age na direção oposta, podendo atenuar as manifestações clínicas das doenças metabólicas. O treinamento físico aeróbio (TFA) tem sido amplamente recomendado para a prevenção e o tratamento de doenças metabólicas, inclusive da DHGNA, e parte das respostas benéficas podem estar associadas com a melhora do metabolismo oxidativo. Assim, o objetivo desse estudo foi investigar se a prevenção da DHGNA por meio do TFA é mediada pela melhora do metabolismo hepático associada à ativação da alça do SRA ECA2/ Ang1-7/ Mas. Para isso, camundongos C57BL/6 foram separados em grupos (n=10/grupo) sedentários (SED) alimentados com dieta normocalórica (NO) ou de cafeteria (CAF) (SED-NO e SED-CAF, respectivamente) e submetidos ao TFA alimentados com dieta NO ou CAF (TF-NO e TF-CAF, respectivamente). O grupo SED-CAF apresentou maior ganho de peso corporal, conteúdo de lipídios e de IL-6 no fígado, e o TFA previniu esses aumentos no grupo TF-CAF. Não houve diferença na concentração sérica das enzimas ALT e AST, na expressão de genes relacionados com o metabolismo lipídico e na expressão das proteínas AMPK, PGC1- , SIRT-1, ACC e receptor Mas no fígado. Os grupos TF-NO e TF-CAF apresentaram maior atividade da ECA2 no soro comparados ao SED-NO e SED-CAF, porém a atividade da ECA2 e o conteúdo do peptídio Ang1-7 não foram diferentes entre os grupos. O TF-NO apresentou menor atividade da enzima ECA no fígado comparado ao grupo TF-CAF. Coletivamente, os dados obtidos permitem afirmar que o TFA preveniu a DHGNA evidenciado pelo menor conteúdo de lipídios e citocina pró-inflamatória IL-6, no entanto, essa resposta foi independente de mudanças na expressão de genes e de proteínas reguladoras do metabolismo hepático associada à alça do SRA ECA2/ Ang1-7/ Mas / Non-alcoholic fatty liver disease (NAFLD) consists in the morphophysiological alteration of the liver due to the accumulation of lipids in the hepatocytes. The development of NAFLD may be associated with obesity and type 2 diabetes, and one of the possible mediating mechanisms is the hyperactivity of the ACE/Ang II/AT1 axis of the renin angiotensin system (RAS). On the other hand, evidence have shown that the activation of the ACE2/Ang 1-7/Mas have opposite effect, being able to attenuate clinical manifestations of the metabolic diseases. Aerobic physical training (APT) has been widely recommended for the prevention and treatment of metabolic diseases, including NAFLD, and some of the beneficial responses may be associated with improved oxidative metabolism. Thus, the aim of this study was to investigate whether the prevention of NAFLD by APT is mediated by the improvement of the metabolism associated with the activation of the RAS ACE2/Ang1-7/Mas axis. For this, C57BL/6 mice were separated into sedentary groups (SED) fed normocaloric (NO) or cafeteria (CAF) diet (SED-NO and SED-CAF, respectively) and trained with APT fed NO or CAF diet (TF-NO and TF-CAF, respectively). The SED-CAF group presented higher body weight gain, lipid and IL-6 content in the liver, and APT prevented these increases in the TF-CAF group. No differences were observed in the concentration of ALT and AST enzymes, in the expression of genes related to lipid metabolism and in the expression of AMPK, PGC1- , SIRT-1, ACC and receptor Mas in the liver. TF-NO and TF-CAF groups had higher serum ACE2 activity compared to SED-NO and SED-CAF, however ACE2 activity and Ang1-7 content were not different among groups. TF-NO showed lower ACE activity in the liver compared to the TF-CAF group. Collectively, the results showed that APT prevented NAFLD evidenced by the lower lipid content and pro-inflammatory cytokine IL-6, however, this response is independent of changes in gene expression and in hepatic metabolism regulatory proteins expression associated with the SRA ACE2/Ang1-7/Mas axis

Background: Obesity has become one of our times most endemic disease on a global scale and changes to lifestyle is the most cost-effective way to treat patients, when the cost for healthcare related treatment is staggeringly high for obesity and sequela diseases NAFLD, diabetes typ 2, dyslipidaemia and metabolic syndrome.The problem with this remedy is that it requires work and dedication. But changes require hard work, and in this patient group- low compliance, weight gain after treatment, dropping out of programs and small desire to change are the most common problems. Motivational studies report that readiness in obese patients is low and the best way to help patients to move forward is by motivational conversations. The obesity sequela disease NAFLD is an asymptomatic disease it displays no symptoms until very late stages. Therefore it’s a problem to get patients make the patient understand his illness and the seriousness of it. Aim: This literature work was aimed at investigating compliance in lifestyle changes in obese subject and to see if close contact with healthcare staff affected the achieved results. Method: In this literature study, the databases Pubmed, Science Direct, Medline and Sportdiscus were used to find information. Article inclusion criteria were that the articles were not older than 10 years and were in English. Result: Frequent and regular contacts between participants and professional staff provided good results both with regard to weight loss, biochemical response, and the participants' willingness to change. Also it shows that return visits at least every three months will improve weight loss if the participant is motivated to implement a change to lifestyle. Conclusion: Overall, this literature study shows the difficulties with lifestyle changes in people with obesity and sequela NAFLD. Close contacts of the patients with healthcare staff has proven to have a positive impact on treatment compliance, but there are other lifestyle difficulties in these patient groups which hamper compliance.

Ce travail doctoral est centré sur la régénération du foie en conditions normales et au cours de la stéatose hépatique. Ces dernières décennies, de nombreux travaux ont utilisé des modèles d’invalidations géniques afin d’identifier les acteurs important au cours de la régénération hépatique. Dans ce contexte, il avait été observé que des animaux dont le signal de l’hormone de croissance était inhibé présentait un défaut majeur de prolifération hépatocytaire après hépatectomie. Dans un premier temps, notre laboratoire s’est donc intéressé à comprendre comment l’hormone de croissance contrôle la régénération hépatique au niveau moléculaire. Pour cela, nous avons pratiqué des hépatectomies sur des animaux dépourvus en récepteur de l’hormone de croissance (GHrKO). Nous avons ainsi montré que l’hormone de croissance jouait un rôle majeur au cours de la régénération hépatique en contrôlant l’expression d’EGFR ainsi que l’activation de Erk1/2. Dans un second temps, je me suis intéressée à une situation pathologique associée à une dérégulation de la voie de l’hormone de croissance : la stéatose hépatique. De façon intéressante, de nombreux modèles murins de stéatose hépatique présentent également une inhibition importante de la prolifération après hépatectomie partielle. Chez l’Homme, cette maladie (NAFLD pour Non alcoholic fatty liver disease) représente un facteur de risque lors de transplantations hépatiques et de résections majeures du foie. Grâce à l’analyse quantifiée de plusieurs paramètres issue de biopsies de patients obèses, nous avons montré l’existence d’une forte corrélation entre stéatose hépatique et diminution de l’expression de l’EGFR sur l’Homme. Nous avons également pratiqué des hépatectomies sur deux modèles de stéatose, l’un génétique (ob/ob) l’autre induit par un régime déficient en méthionine et choline (MCD). Les cinétiques de régénération post hépatectomie nous ont permis de confirmer un défaut de régénération hépatique chez les souris ob/ob et MCD. D’autre part, l’étude de ces modèles de stéatose nous a amenés à valider la dérégulation de la voie de l’hormone de croissance et la diminution transcriptionnelle de l’EGFR avant et après hépatectomie partielle. En parallèle, nous avons souligné l’implication de la voie inhibitrice de prolifération TGF-β, dans l’altération de la prolifération hépatocytaire des animaux ob/ob. En effet, de nombreux acteurs de cette voie sont surexprimés après l’hépatectomie partielle, participant certainement au défaut de régénération plus drastique observé sur ce modèle. Pour finir, nous avons également montré que l’injection sur une courte période d’hormone de croissance sur les animaux ob/ob restaure la prolifération hépatocytaire post hépatectomie. Ce sauvetage phénotypique est associé à une réexpression transcriptionnelle et protéique de l’EGFR. A terme, ces travaux nous amènent à proposer que la dérégulation de l’axe hormone de croissance/EGFR représente un mécanisme général associé à la stéatose hépatique et responsable du défaut de régénération du foie lié à cette maladie. / This doctoral work focused on liver regeneration in physiological conditions and during steatosis. These last decades, several studies used gene invalidation models to identify important actors during the liver regeneration. In this context, it had been observed that animals displaying a defect of growth hormone pathway had a drastic defect of liver regeneration after partial hepatectomy. Initially, we started this work by focusing on understanding how growth hormone controls liver regeneration at the molecular level. To do so, we performed partial hepatectomies on animals deleted for the growth hormone receptor gene (GHrKO). These results showed that growth hormone plays a central role in the control of liver regeneration through the expression of EGFR and the activation of Erk1/2. Secondly, we focused our attention on a pathological situation showing a defect of growth hormone signaling : hepatic steatosis. Interestingly, many mice models of hepatic steatosis also present a drastic inhibition of hepatocytes proliferation after partial hepatectomy. In Humans, non-alcoholic fatty liver disease (NAFLD) represents an important risk factor regarding liver transplantations and resections. Through quantified analysis of several parameters from obese patient biopsies, we showed the existence of a strong correlation between hepatic steatosis and decrease in EGFR expression on humans. We also performed partial hepatectomies on two models of hepatic steatosis, one being genetic (ob/ob) and the other one being induced by a methionine choline deficient diet (MCD). Kinetics of regeneration post hepatectomy led us to confirm the defect of liver regeneration in on ob/ob and MCD mice. Moreover, the study of these steatotic models allowed us to corroborate the downregulation of the growth hormone signaling and the transcriptional decrease of EGFR expression. We also underlight the importance of TGF-β, a signaling pathway inhibiting proliferation, in the liver regeneration defect observed in ob/ob mice. Indeed, many members of this pathway have been found to be upregulated after partial hepatectomy, possibly being involved in the drastic regeneration defect observed in ob/ob mice. To finish, we also showed that growth hormone injections on a small period of time in ob/ob mice were capable of rescuing hepatocyte proliferation post hepatectomy. This phenotypic rescue was associated with a reexpression of EGFR at the transcription and protein level. This work led us to propose that the defect of the growth hormone/EGFR pathway represents a general mechanism associated with hepatic steatosis and is responsible for the liver regeneration defect linked to this disease.

Il est estimé aujourd’hui que plus de 350 médicaments peuvent induire des lésions hépatiques entraînant différentes manifestations cliniques telles qu’une hépatite cytolytique, une stéatose voire une cirrhose. Bon nombre de médicaments hépatotoxiques induisent un dysfonctionnement mitochondrial. Cependant, les mécanismes induisant de tels effets délétères ne sont pas tous élucidés, en particulier ceux concernant l’ADN mitochondrial (ADNmt) et son homéostasie, qui ne sont pas souvent explorés. De plus, il existe peu d’informations concernant l’hépatotoxicité médicamenteuse dans un contexte de stéatose induite par l’obésité. Ainsi, l’objectif de ce travail a été tout d’abord de mettre au point un modèle de stéatose dans les cellules de la lignée hépatocytaire humaine HepaRG afin d’étudier ensuite, les effets de neuf médicaments hépatotoxiques et vraisemblablement mitochondriotoxiques – l’amiodarone, l’atorvastatine, la carbamazépine, l’imipramine, la lovastatine, la perhexiline, le ritonavir, la terbinafine et la troglitazone – sur l’homéostasie de l’ADNmt dans un contexte ou non de stéatose. En utilisant des concentrations peu ou non cytotoxiques, nous avons trouvé que parmi les neuf médicaments étudiés, le ritonavir et l’imipramine ont induit des effets mitochondriaux suggérant une altération de la traduction mitochondriale. De façon notable, la toxicité du ritonavir était plus importante dans les cellules non-stéatosées. De plus, aucun des neuf médicaments n’a induit de diminution des quantités d’ADNmt. Cependant, les quantités accrues d’ADNmt ont été retrouvées avec six des neuf médicaments, et notamment dans les cellules non-stéatosées. Cela était par ailleurs accompagné d’une modulation de l’expression des différents facteurs impliqués dans la biogenèse mitochondriale (PGC-1α, PGC-1β, AMPK, etc.). Ainsi, ces données laissent supposer qu’une altération de la traduction mitochondriale peut ne pas être une événement rare et que l’augmentation des quantités d’ADNmt et la modulation de la biogenèse mitochondriale pourraient être une réponse adaptative fréquente à des altérations mitochondriales pouvant être amoindrie par la stéatose. / It is currently estimated that more than 350 drugs can induce liver injury with different clinical presentations such as hepatic cytolysis, steatosis, even cirrhosis. Many hepatotoxic drugs can induce mitochondrial damage and dysfunction. However, not all mechanisms that lead to such deleterious effects are clarified, especially those concerning mitochondrial DNA (mtDNA) and its homeostasis, which are not often investigated. Moreover, there is little information regarding the impact of non alcoholic fatty liver disease (NAFLD) on drug-induced liver injury. Thus, the aim of this work was, first of all, to develop a model of NAFLD in the hepatic cell line HepaRG in order to study further effects of nine hepatotoxic and presumably mitochondriotoxic drugs – amiodarone, atorvastatin, carbamazepine, imipramine, lovastatin, perhexiline, ritonavir, terbinafine and troglitazone –, on mtDNA homeostasis in the context of NAFLD or not. By using drug concentrations that did not induce major cytotoxicity, we found that, among the nine drugs, studied, ritonavir and imipramine induced mitochondrial effects suggesting alteration of mtDNA translation. Notably, ritonavir toxicity was stronger in non-steatotic cells. Furthermore, none of the nine drugs decreased mtDNA levels. However, increased mtDNA was observed with six drugs, especially in non-steatotic cells. This result was also accompanied by a modulation of the expression of various factors involved in mitochondrial biogenesis (e.g. PGC-1α, PGC-1β, AMPK).Therefore, this data suggests that drug-induced impairment of mtDNA translation may not be a rare event and increased mtDNA levels and modulation of mitochondrial biogenesis could be a frequent adaptive response to mitochondrial impairments, which could be dampened by steatosis.

To compare ultrasound-based acoustic structure quantification (ASQ) with established non-invasive techniques for grading and staging fatty liver disease.

La stéatose hépatique non alcoolique (SHNA) est maintenant une des principales causes de cancer du foie. Cependant, les mécanismes physiopathologiques contribuant à son développement ou à la progression de la maladie sont peu connus. Il a été démontré que le niveau d’expression du coactivateur transcriptionnel PGC-1α est inversement proportionnel avec la sévérité de la stéatose hépatique le stress oxydatif et la résistance à l’insuline dans les foies de souris. Chez l’humain, on observe aussi une diminution de PGC-1α dans les foies de patients atteints de SHNA. De plus, il a été démontré que les souris avec une réduction de 50% des niveaux hépatique de PGC-1α mène à une sensibilité à l’insuline et à une tolérance au glucose altérée dans les tissus périphériques. Ces découvertes suggèrent qu’en plus d’être associés au développement de la SHNA, les niveaux hépatiques de PGC-1α altèrent l’expression de facteurs sécrétoires du foie afin d’influencer la régulation métabolique de tout le corps. Nous proposons qu’une réduction de l’expression de PGC-1α dans le foie influence les protéines sécrétées par le foie en situation de stress métabolique, révélant l’importance de PGC-1α dans la réponse adaptative du foie. L’analyse du sécrétome hépatique effectuée par spectrométrie de masse sur le milieu conditionné d’hépatocytes primaires a identifié SERPINA3N, une protéine sécrétée, dont les niveaux corrèlent avec les niveaux hépatiques de PGC-1α et sont influencés par la diète obésogène. Dans ce projet, les niveaux sanguins de cette protéine ont été quantifiés par western blot chez des souris mâles et femelles, sauvages ou hétérozygotes pour PGC-1α dans le foie et nourris avec une diète control ou riche en gras et en fructose. Nos résultats démontrent que les niveaux circulatoires de SERPINA3N augmentent avec la diète et corrèlent avec les niveaux hépatiques de PGC-1α de manière dépendante à la diète et le sexe. De plus, les niveaux sanguins de SERPINA3N diminuent avec la progression de la maladie. L’expression hépatique de SERPINA3N est grandement influencée par les niveaux de PGC-1α, mais indépendamment du facteur transcriptionnel NF-κB. Nous avons montré que les glucocorticoïdes augmentent les niveaux protéiques et circulatoires de SERPINA3N dans les hépatocytes primaires. De plus, cette augmentation par les glucocorticoïdes est influencée par les niveaux de PGC-1α. Ces résultats révèlent une nouvelle interaction entre PGC-1α et le récepteur des glucocorticoïdes sur l’expression hépatique et la sécrétion de SERPINA3N. Pour conclure, l’identification de protéines circulatoires régulées par PGC-1α nous aidera à mieux comprendre comment la perte d’expression de PGC-1α dans le foie affecte le métabolisme de tout le corps dans le contexte de la SHNA. / Non-alcoholic fatty liver disease (NAFLD) is becoming a serious public health problem and is now one of the leading causes of liver cancer. Although NAFLD is known to be associated with obesity, insulin resistance, metabolic syndrome and type II diabetes, the mechanisms contributing to its development are not fully understood. It is shown that hepatic PGC-1α levels correlate negatively with NAFLD development, oxidative liver damage and hepatic insulin resistance in murine models. In humans, decrease PGC-1α expression in NAFLD and NASH patients. Moreover, liver-specific PGC-1α reduction in mice also disrupts glucose tolerance and insulin sensitivity in muscle and adipose tissue, likely due to altered secretion of hepatic hormones. These findings suggest that in addition to contributing to NAFLD development, the hepatic disruption of PGC-1α alters the liver secretome, thereby influencing the whole-body energy metabolism. We hypothesize that decreased expression of PGC-1α in the liver alters the expression of hepatokines under metabolic challenges, revealing a potential novel role for PGC-1α in the adaptive response of the liver. The hepatocyte-specific secretome was analyzed by mass spectrometry (iTRAQ) in conditioned media from primary hepatocytes. We identified SERPINA3N, a secreted protein whose secreted levels correlated with hepatic PGC-1α levels in a diet-dependent manner. This hepatokine was measured in serum from male, female, wildtype and liver-specific PGC-1α heterozygote mice fed chow or high-fat, high-fructose diet using western blot. SERPINA3N circulating levels increased with the western diet and correlated with hepatic PGC-1α levels in a diet and sex-dependent manner. Its serum levels decreased with the progression of the disease. The hepatic SERPINA3N expression was greatly influenced by PGC-1α levels independently of NF-κB transcription factor. We showed that glucocorticoids increased SERPINA3N protein and secreted levels in primary hepatocytes. This increase was influenced by PGC-1α levels, revealing a novel interaction of PGC-1α and the glucocorticoid receptor on SERPINA3N expression and secretion. In conclusion, this project reveals a novel impact of hepatic PGC-1α levels on the liver secretome during NAFLD development. This work will provide insights on the role of hepatic PGC-1α levels on the regulation of hepatokines and how it influences the whole-body energy homeostasis in a context of NAFLD.

Spinal muscular atrophy (SMA) is a devastating recessive neurological disorder thought to be affecting primarily the motor neurons. As such, paralysis, motor weakness and death ensue. While SMA is most commonly seen in infants and children, it can span all ages. Its genetic etiology revolves around the homozygous deletion or mutation of the SMN1 gene, whose product (SMN protein) has critical and ubiquitous roles in mRNA splicing, amongst various other functions in mRNA metabolism. As such, SMN depletion in other non-neuronal cells type is likely to have physiological repercussions, and perhaps modulate the SMA phenotype. Herein, we identify the molecular pathways of atrophy in skeletal and cardiac muscle of two mouse models of SMA and their therapeutic modulation via the histone deacetylase inhibitor trichostatin A. We also identify dramatic changes in immune organs in mouse models of SMA, which could impact susceptibility to infections. Furthermore, we establish the presence of important defects in fatty acid homeostasis in the liver and plasma seen in both mouse models and SMA patients. Finally, we provide the first mild mouse model of SMA that reliably reproduces canonical features of SMA, permitting aging studies. This model presents with a prominent myopathic phenotype prior to motor neuron death, without extra-neuronal involvement during the course of its lifespan. Overall, our work shows multiple potentially clinically relevant defects in extra-neuronal organs, provides ways to abrogate them and provides a framework to study them over the course of aging.

No description available.

Thesis (MSCMedSc)--Stellenbosch University, 2012. / Includes bibliography / ENGLISH ABSTRACT: Non-alcoholic fatty liver disease (NAFLD) is the most prevalent chronic liver disease in the world. The disease spectrum of NAFLD extends from steatosis (types 1,2) to non-alcoholic steatohepatitis (NASH) with inflammation (types 3,4). The aims of the study were 1) to analytically validate high-throughput real time polymerase chain reaction (RT-PCR) assays for three selected single nucleotide polymorphisms (SNPs), FTO rs9939609 (intron 1 T>A), TNF-α rs1800629 (-308 G>A) and PPARγ rs1801282 (Pro12Ala, 34 C>G), and 2) to perform genotype-phenotype association studies in relation to biochemical abnormalities, disease severity and age of onset. A total of 119 patients with fatty liver identified on ultrasound, including 88 histologically confirmed NAFLD patients, and 166 control individuals were genotyped for the three selected SNPs. RT-PCR validated against direct sequencing as the gold standard was used for detection of genetic variation. All three SNPs were in Hardy Weinberg equilibrium in the study population, except for a deviation in genotype distribution detected for PPARγ rs1801282 in the NAFLD patient subgroup (p<0.001). After adjustment for age and gender, the risk-associated FTO rs9939609 A-allele was detected at a significantly higher frequency in the Caucasian compared with Coloured patients (p=0.005). The opposite was detected for the risk-associated TNF-α rs1800629 A-allele, which occurred at a significantly higher frequency in the Coloured compared with Caucasian NAFLD patients (p=0.034). The onset of fatty liver disease symptoms was on average 5 years younger in the presence of each risk-associated TNF-α rs1800629 A-allele (p=0.028). When considered in the context of an inferred genotype risk score ranging from 0-6, disease onset occurred on average 3 years earlier (p=0.008) in the presence of each risk-associated FTO A-allele, TNF-α A-allele or PPARγ C-allele. After adjustment for age, gender and race, no differences in genotype distribution or allele frequencies were observed between histologically confirmed NAFLD (types 1,2) and NASH (types 3,4) patients, while the minor allele frequency for the TNF-α rs1800629 was significantly higher in the total NAFLD (types 1-4) (p=0.047) as well as NASH subgroup (NAFLD types 3,4) (p=0.030) compared with obese patients without a histologically confirmed NAFLD diagnosis. A significant correlation was furthermore observed between the number of TNF-α rs1800629 A-alleles and increasing CRP levels (p=0.029), with a favourable reduced effect in the presence of low- to moderate alcohol intake. The average waist circumference of physically active NAFLD patients was 12% lower than in physically inactive patients (p=0.004). In view of the results presented in this study, the inclusion of the selected SNPs, and in particular the pro-inflammatory TNF-α rs1800629 polymorphism, may be considered as part of a comprehensive cardiovascular risk evaluation of NAFLD patients. Ultimately, early detection of patients with fatty liver disease symptoms and effective intervention based on the underlying disease mechanism may prevent progression from NAFLD to NASH, shown to be an independent risk factor for cardiovascular diseases. / AFRIKAANSE OPSOMMING: Nie-alkoholiese lewervervetting (NALV) is die mees algemene kroniese lewersiekte in die wêreld. Die siektespektrum van NALV strek van steatose (vervette lewer tipes 1,2) tot steatohepatitis met inflammasie (NASH tipes 3,4). Die doel van die studie was 1) om analities die hoë omset polimerase kettingreaksie (RT-PKR) metode te valideer vir die geselekteerde enkel nukleotied polimorfismes (ENPs) FTO rs9939609 (intron 1 T>A), TNF-α rs1800629 (-308 G>A) en PPARγ rs1801282 (Pro12Ala, 34 C>G), en 2) om genotipe-fenotipe assosiasie studies uit te voer ten opsigte van relevante biochemiese abnormaliteite, graad van die siekte en aanvangsouderdom. ’n Totaal van 119 pasiënte met vervette lewers is geïdentifiseer met behulp van ultraklank, insluited 88 histologies-bevestigde NALV pasiënte, en 166 kontrole individue. Hierdie pasiënte is gegenotipeer vir die 3 geselekteerde ENP’s. RT-PKR gevalideer met direkte DNA volgorde bepaling as die goue standaard, is gebruik vir opsporing van genetiese variasie. Al die ENP’s was in Hardy Weinberg ekwilibrium in die studie populasie, behalwe vir ’n afwyking in genotipe verspreiding waargeneem vir PPARγ in die NALV subgroep (p<0.001). Nadat aanpassings gemaak is vir ouderdom en geslag, is die risiko-geassosieerde FTO rs9939609 A-alleel waargeneem teen ’n betekenisvol hoër frekwensie in die Kaukasiese pasiënte in vergelyking met Kleurling pasiënte (p=0.005). Die teenoorgestelde is waargeneem vir die risiko-geassosieerde TNF-α rs1800629 A-alleel wat voorgekom het teen ’n betekenisvol hoër frekwensie in die Kleurling NALV pasiënte, in vergelyking met Kaukasiese NALV pasiënte (p=0.034). Die aanvang van NALV was gemiddeld 5 jaar vroeër in die teenwoordigheid van elke risiko-geassosieerde TNF-α rs1800629 A-alleel (p=0.028). Met inagneming van ’n genotipe risiko telling tussen 0–6, het aanvang van siekte gemiddeld 3 jaar vroeër voorgekom (p=0.008) in die teenwoordigheid van elke toenemende risiko-geassosieerde FTO A-alleel, TNF-α A-alleel en PPARγ C-alleel. Nadat aanpassings gemaak is vir ouderdom, geslag en ras, is geen verskille waargeneem in genotipe verspreiding of alleel frekwensies tussen histologies bevestigde NALV (tipes 1,2) en NASH (tipes 3,4) pasiënte nie, terwyl die minor alleel telling vir die TNF-α rs1800629 betekenisvol hoër was in die totale NALV (tipes 1–4) (p=0.047) asook die NASH subgroep (NALV tipes 3,4) (p=0.03) in vergelyking met vetsugtige pasiënte sonder ’n histologies bevestigde diagnose. ‘n Statisties beteknisvolle korrelasie is verder waargeneem tussen die aantal TNF-α rs1800629 A-allele en toenemende CRP vlakke (p=0.029), met n gunstige verlaagde effek in die teenwoordigheid van lae alcohol gebruik. Die gemiddelde middellyf-omtrek van fisies aktiewe NALV pasiënte was 12% minder as fisies onaktiewe pasiente (p=0.004). Na aanleiding van die resultate van hierdie studie behoort insluiting van geselekteerde ENP’s, en in besonder die pro-inflammatoriese TNF-α rs1800629 polimorfisme, as deel van ’n omvattende kardiovaskulere risiko evaluasie oorweeg te word. Aan die einde van die dag mag vroeë identifikasie van NALV pasiente en effektieve intervensie gebasseer op die onderliggende siekte meganisme, vordering tot NASH verhoed wat getoon is om ’n onafhanklike risiko faktor vir kardiovaskulêre siekte te wees. / Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology