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Yes / Abstract Objective Omega-3 polyunsaturated fatty acids (PUFAs) have anticolorectal cancer (CRC) activity. The intestinal microbiota has been implicated in colorectal carcinogenesis. Dietary omega-3 PUFAs alter the mouse intestinal microbiome compatible with antineoplastic activity. Therefore, we investigated the effect of omega-3 PUFA supplements on the faecal microbiome in middle-aged, healthy volunteers (n=22). Design A randomised, open-label, cross-over trial of 8 weeks’ treatment with 4 g mixed eicosapentaenoic acid/docosahexaenoic acid in two formulations (soft-gel capsules and Smartfish drinks), separated by a 12-week ‘washout’ period. Faecal samples were collected at five time-points for microbiome analysis by 16S ribosomal RNA PCR and Illumina MiSeq sequencing. Red blood cell (RBC) fatty acid analysis was performed by liquid chromatography tandem mass spectrometry. Results Both omega-3 PUFA formulations induced similar changes in RBC fatty acid content, except that drinks were associated with a larger, and more prolonged, decrease in omega-6 PUFA arachidonic acid than the capsule intervention (p=0.02). There were no significant changes in α or β diversity, or phyla composition, associated with omega-3 PUFA supplementation. However, a reversible increased abundance of several genera, including Bifidobacterium, Roseburia and Lactobacillus was observed with one or both omega-3 PUFA interventions. Microbiome changes did not correlate with RBC omega-3 PUFA incorporation or development of omega-3 PUFA-induced diarrhoea. There were no treatment order effects. Conclusion Omega-3 PUFA supplementation induces a reversible increase in several short-chain fatty acid-producing bacteria, independently of the method of administration. There is no simple relationship between the intestinal microbiome and systemic omega-3 PUFA exposure. / NIHR/EME Yorkshire Cancer Research (YCR)

Le tractus digestif et le foie interagissent continuellement, non seulement à travers les connexions anatomiques, mais également par des liens physiologiques/fonctionnels. Le déséquilibre de l’axe intestin-foie apparait de plus en plus comme un facteur primordial dans les désordres cardiométaboliques, à savoir l’obésité, le syndrome métabolique, le diabète de type 2 et la stéatose hépatique non alcoolique (NAFLD), pour lesquels la prévalence demeure alarmante, les mécanismes moléculaires encore méconnus, et les traitements peu efficaces. L’hypothèse centrale du présent projet de recherche est que la combinaison d’anomalies génétiques et nutritionnelles affecte la sensibilité de l’insuline intestinale, ce qui conduit à une surproduction des chylomicrons, à une dyslipidémie, une insulinorésistance systémique et des répercussions sur le foie. Dans cet agencement, le foie développe une NAFLD progressive, impliquant plusieurs sentiers métaboliques intrinsèques et des mécanismes comprenant le stress oxydatif, l’inflammation et l’insulinorésistance. En revanche, des nutriments, comme les acides gras polyinsaturés (AGPI) n-3, peuvent présenter des effets bénéfiques en ciblant plusieurs circuits pathogéniques. L’objectif central de cette thèse consiste à : (i) Démontrer que des gènes codant pour les protéines intestinales clés associées au transport des lipides, comme c’est le cas du Sar1b GTPase, peuvent interagir avec l’environnement nutritionnel pour produire l’obésité et des dérangements cardiométaboliques, incluant la NAFLD ; (ii) Explorer les mécanismes hépatiques sous-jacents à la NAFLD; et (iii) Identifier les effets et les cibles thérapeutiques des AGPI n-3 sur la NAFLD. Ces objectifs seront soutenus par une prospection de la littérature scientifique disponible dans les champs du syndrome métabolique et de la NAFLD afin d’en disséquer les forces et les faiblesses au bénéfice de la communauté scientifique. À ces fins, nous avons utilisé des modèles animaux et cellulaires manipulés génétiquement, des animaux exposés de façon chronique à des diètes riches en lipides, des spécimens de tissus hépatiques obtenus durant la chirurgie bariatrique d’obèses morbides, et une cohorte d’adolescents obèses souffrant de NAFLD et qui seront traités avec les AGPI n-3. L’ensemble de nos expériences ont soutenu nos hypothèses et ont mis en évidence les concepts et mécanismes suivants : (i) L’abondance d’un gène crucial (notamment Sar1b GTPase) au niveau de l’intestin, en synergie avec une alimentation obésogène, perturbe l’homéostasie locale et mène à des dérangements cardiométaboliques, défiant même l’axe intestin-foie ; (ii) Les causes développementales de la NAFLD comprennent les dérangements du métabolisme des acides gras, du statut redox et inflammatoire, de la sensibilité à l’insuline, des sentiers métaboliques (lipogenèse, β-oxydation, gluconéogenèse) et de l’expression des facteurs de transcription; et (iii) Les AGPI n-3 représentent un robuste arsenal thérapeutique des dérangements cardiométaboliques, notamment la NAFLD, en agissant sur plusieurs cibles pathogéniques. Globalement, nos résultats montrent le rôle indéniable de l’intestin comme organe insulino-sensible interagissant de près avec les aliments et capable de déclencher des troubles métaboliques. Plusieurs mécanismes gouvernant les désordres métaboliques ont été dévoilés par nos travaux. En outre, nos études cliniques ont pointé la force thérapeutique des AGPI n-3 qui interviennent dans de nombreux processus de régulation métaboliques et notamment dans le stress oxydatif et l’inflammation. / The digestive tract and liver interact continuously, not only through anatomical connections, but also through physiological / functional links. The imbalance of the intestine-liver axis is increasingly emerging as a key factor in cardiometabolic disorders (CMD), namely obesity, metabolic syndrome, type 2 diabetes, and non alcoholic fatty liver disease (NAFLD), for which prevalence remains alarmingly high, molecular mechanisms are poorly understood, and treatments are largely inefficient. The central hypothesis of this research project is that the combination of genetic and nutritional abnormalities affect intestinal insulin sensitivity, leading to overproduction of chylomicrons, dyslipidemia, systemic insulin resistance and dysregulated intestine-liver axis. In this situation, the liver develops progressive NAFLD, implicating several intrinsic metabolic pathways and mechanisms, including oxidative stress, inflammation and insulin resistance. In contrast, functional foods, such as omega-3 polyunsaturated fatty acids (n-3 PUFA), may have beneficial effects by targeting several pathogenic pathways. The central objective of this thesis is to: (i) Demonstrate that genes coding for key intestinal proteins associated with lipid transport, as is the case with Sar1b GTPase, can interact with the nutritional environment to produce obesity and CMD, including hepatic steatosis; (ii) explore the mechanisms underlying NAFLD; and (iii) identify the effects and therapeutic targets of n-3 PUFA. These objectives will be supported by a critical review on metabolic syndrome and NAFLD in order to dissect their strengths and weaknesses for the benefit of the scientific community. For these purposes, we used genetically engineered animal and cell models, chronic exposure of animals to high-fat diets, liver tissue specimens obtained during bariatric surgery of morbidly obese patients, and treatment of obese NAFLD adolescents with n-3 PUFA. All of our experiments supported our hypotheses and highlighted the following concepts and mechanisms: (i) The abundance of a crucial gene (notably Sar1b GTPase) in the intestine, in synergy with an obesogenic diet, disrupts local homeostasis and leads to CMD, challenging even the intestine-liver axis; (ii) Developmental causes of NAFLD include disturbances of fatty acid metabolism, redox and inflammatory status, insulin sensitivity, metabolic pathways (lipogenesis, β-oxidation, gluconeogenesis), and expression of transcription factors; and (iii) n-3 PUFA represent a robust therapeutic arsenal of CMD, including NAFLD, by acting on several pathogenic targets. Overall, our results show the undeniable role of the intestine, as an insulin-sensitive organ, interacting closely with obesogenic food, and capable of triggering CMD, including perturbations of the intestine-liver axis. Several mechanisms governing metabolic disorders have been unveiled by our work. In addition, our clinical studies have pointed to the therapeutic potential of n-3 PUFA involved in many regulatory processes, including oxidative stress and inflammation.

Une mort cellulaire par apoptose impliquant un processus inflammatoire est observée dans le système limbique, suite à un infarctus du myocarde. Les oméga-3 et ses métabolites, en plus de leurs propriétés bénéfiques pour le système cardiovasculaire, réduisent l’inflammation, contrairement aux oméga-6 qui sont plus pro-inflammatoires. Comme le métabolisme de ces deux acides gras essentiels impliquent les mêmes enzymes, le ratio alimentaire oméga-3/6 aurait donc des impacts importants sur l'état inflammatoire et ainsi indirectement sur l'apoptose. Conséquemment, cette étude a pour but d'évaluer l'effet de différents ratios oméga-3/6 sur la taille de l’infarctus, l’inflammation et l’apoptose dans le système limbique suite à un infarctus du myocarde. Des rats Sprague-Dawley ont été aléatoirement distribués dans trois groupes contenant des ratios 1:1, 1:5 et 5:1 oméga-3/6. Ils ont été nourris pendant 2 semaines, suivie d’une occlusion de l’artère coronaire gauche descendante pendant 40 minutes et d’une période de reperfusion (15 min et 24 h). De hauts ratios d’oméga-3 (5:1 et 1:1) diminuent significativement la taille de l’infarctus de 32 % et augmentent l’activité d’Akt, impliquée dans la voie cardioprotectrice RISK, comparativement au ratio 1:5. Ils diminuent aussi la concentration plasmatique de TNF-D. Dans le système limbique, l’activité de la caspase-3 est augmentée dans la région CA1, après 15 min, et dans les régions du CA1 et du gyrus dentelé (Gd), après 24 h, avec la diète 1:5 en comparaison aux diètes 1:1 et 5:1. L’activité enzymatique de la caspase-8 est augmentée dans le Gd, alors que dans le CA1, il y a une activité plus importante de la caspase-9 aux temps de reperfusion étudiés. Conclusion: Les diètes élevées en oméga-3/oméga-6 réduisent la taille de l'infarctus, l’inflammation et diminuent l’apoptose dans le système limbique après un infarctus du myocarde. / Apoptosis occurs in the limbic system by a mechanism involving inflammation after a myocardial infarction. Omega-3 are essential fatty acids known for their benefits in cardiovascular health and to reduce inflammation, whereas, omega-6 and their metabolites are more inflammatory. Since these two essential fatty acids are metabolized by the same group of enzymes, the resulting competition would affect the inflammation and thus indirectly the apoptosis. Therefore, the aim of this study is to evaluate the effect of different omega-3/6 ratios on the infarct size, inflammation and apoptosis in the limbic system after myocardial infarction. Male Sprague-Dawley rats were divided randomly between three diet groups containing 1:5, 1:1 and 5:1 omega-3/6 ratios. Rats were fed for two weeks followed by a left anterior descending coronary occlusion for 40 min and by reperfusion time (15 min or 24 h). Infarct size was significantly reduced by 32% and Akt activity, implicated in RISK cardioprotection pathway, enhanced by the 5:1 and 1:1 in comparison to the 1:5 diet ratios. They also reduced TNF-D plasma concentration. In the limbic system, caspase-3 enzymatic activity was doubled in CA1 after 15 min and in CA1 and dentate gyrus (Dg), after 24 h, in the 1:5 compared to 1:1 and 5:1 groups. Caspase-8 enzymatic activity was increased in Dg and capase-9 was enhanced in the CA1 at both reperfusion time in the 1:5 against the 1:1 and 5:1 groups. Conclusion: High omega-3 dietary ratio helps to reduce infarct size, inflammation and to prevent apoptosis in the limbic system.

L’athérosclérose est la principale cause d’infarctus du myocarde, de mort subite d’origine cardiaque, d’accidents vasculaires cérébraux et d’ischémie des membres inférieurs. Celle-ci cause près de la moitié des décès dans les pays industrialisés. Lorsque les obstructions artérielles athérosclérotiques sont tellement importantes que les techniques de revascularisation directe ne peuvent être effectuées avec succès, la sévérité de l’ischémie tissulaire résiduelle dépendra de l’habilité de l’organisme à développer spontanément de nouveaux vaisseaux sanguins (néovascularisation). La néovascularisation postnatale est le résultat de deux phénomènes : la formation de nouveaux vaisseaux à partir de la vasculature existante (angiogenèse) et la formation de vaisseaux à partir de cellules souches progénitrices (vasculogenèse). Notre laboratoire a démontré que plusieurs facteurs de risque associés aux maladies cardiovasculaires (tabagisme, vieillissement, hypercholestérolémie) diminuaient également la réponse angiogénique suite à une ischémie. Cependant, les mécanismes précis impliqués dans cette physiopathologie sont encore inconnus. Un point commun à tous ces facteurs de risque cardiovasculaire est l’augmentation du stress oxydant. Ainsi, le présent ouvrage visait à élucider l’influence de différents facteurs de risque cardiovasculaire et du stress oxydant sur la néovascularisation. Nos résultats démontrent que l’exposition à la fumée de cigarette et le vieillissement sont associés à une diminution de la néovascularisation en réponse à l’ischémie, et que ceci est au moins en partie causé par une augmentation du stress oxydant. De plus, nous démontrons que les acides gras dérivés de la diète peuvent affecter la réponse à l’ischémie tissulaire. La première étude du projet de recherche visait à évaluer l’impact de l’exposition à la fumée de cigarette sur la néovascularisation post-ischémique, et l’effet d’une thérapie antioxydante. L’exposition à la fumée de cigarette a été associée à une réduction significative de la récupération du flot sanguin et de la densité des vaisseaux dans les muscles ischémiques. Cependant, une récupération complète de la néovascularisation a été démontrée chez les souris exposées à la fumée de cigarette et traitées au probucol ou aux vitamines antioxydantes. Nous avons démontré qu’une thérapie antioxydante administrée aux souris exposées à la fumée de cigarette était associée à une réduction significative des niveaux de stress oxydant dans le plasma et dans les muscles ischémiques. De plus, les cellules endothéliales progénitrices (EPCs) exposées à de l’extrait de fumée de cigarette in vitro présentent une diminution significative de leur activité angiogénique (migration, adhésion et incorporation dans les tissus ischémiques) qui a été complètement récupérée par le probucol et les vitamines antioxydantes. La deuxième étude avait pour but d’investiguer le rôle potentiel de la NADPH oxydase (Nox2) pour la modulation de la néovascularisation post-ischémique dans le contexte du vieillissement. Nous avons trouvé que l’expression de la Nox2 est augmentée par le vieillissement dans les muscles ischémiques des souris contrôles. Ceci est associé à une réduction significative de la récupération du flot sanguin après l’ischémie chez les vieilles souris contrôles comparées aux jeunes. Nous avons aussi démontré que la densité des capillaires et des artérioles est significativement réduite dans les muscles ischémiques des animaux vieillissants alors que les niveaux de stress oxydant sont augmentés. La déficience en Nox2 réduit les niveaux de stress oxydant dans les tissus ischémiques et améliore la récupération du flot sanguin et la densité vasculaire chez les animaux vieillissants. Nous avons aussi démontré que l’activité fonctionnelle des EPCs (migration et adhésion à des cellules endothéliales matures) est significativement diminuée chez les souris vieillissantes comparée aux jeunes. Cependant, la déficience en Nox2 est associée à une récupération de l’activité fonctionnelle des EPCs chez les animaux vieillissants. Nous avons également démontré une augmentation pathologique du stress oxydant dans les EPCs isolées d’animaux vieillissants. Cette augmentation de stress oxydant dans les EPCs n’est pas présente chez les animaux déficients en Nox2. La troisième étude du projet de recherche a investigué l’effet des acides gras dérivés de la diète sur la néovascularisation postnatale. Pour ce faire, les souris ont reçu une diète comprenant 20% d’huile de maïs (riche en oméga-6) ou 20% d’huile de poisson (riche en oméga-3). Nos résultats démontrent qu’une diète riche en oméga-3 améliore la néovascularisation post-ischémique au niveau macro-vasculaire, micro-vasculaire et clinique comparée à une diète riche en oméga-6. Cette augmentation de la néovascularisation postnatale est associée à une réduction du ratio cholestérol total/cholestérol HDL dans le sérum et à une amélioration de la voie VEGF/NO dans les tissus ischémiques. De plus, une diète riche en acides gras oméga-3 est associée à une augmentation du nombre d’EPCs au niveau central (moelle osseuse) et périphérique (rate). Nous démontrons aussi que l’activité fonctionnelle des EPCs (migration et incorporation dans des tubules de cellules endothéliales matures) est améliorée et que le niveau de stress oxydant dans les EPCs est réduit par la diète riche en oméga-3. En conclusion, nos études ont permis de déterminer l’impact de différents facteurs de risque cardiovasculaire (tabagisme et vieillissement) et des acides gras dérivés de la diète (oméga-3) sur la néovascularisation post-ischémique. Nous avons aussi identifié plusieurs mécanismes qui sont impliqués dans cette physiopathologie. Globalement, nos études devraient contribuer à mieux comprendre l’effet du tabagisme, du vieillissement, des oméga-3, et du stress oxydant sur l’évolution des maladies vasculaires ischémiques. / Atherosclerosis is the main cause of myocardial infarction, sudden cardiac death, stroke and lower limb ischemia. It is responsible for nearly half of all deaths in industrialized countries. When atherosclerotic arterial obstructions are so important that direct revascularization techniques cannot be successfully performed, the severity of residual tissue ischemia depends on the ability of the organism to spontaneously develop new blood vessels (neovascularization). Postnatal neovascularization is the result of two phenomena: the formation of new bloods vessels from the existing vasculature (angiogenesis) and vessel formation from progenitor cells (vasculogenesis). Our laboratory has demonstrated that several cardiovascular risk factors (smoking, aging, and hypercholesterolemia) also impair the angiogenic response after ischemia. However, the precise mechanisms involved in that pathophysiology are still unknown. A common feature of all the cardiovascular risk factors is increased oxidative stress. Therefore, the purpose of the present work was to elucidate the influence of cardiovascular risk factors and oxidative stress on neovascularization. Our results demonstrate that exposure to cigarette smoke and aging are associated with impaired neovascularization in response to ischemia, and that this is at least in part due to increased oxidative stress. In addition, we demonstrate that fatty acids derived from the diet can modulate the response to tissue ischemia. The first study of the research project evaluated the effect of cigarette smoke exposure on neovascularization in response to ischemia, and the effect of an antioxidant therapy. Exposure to cigarette smoke was associated with a significant reduction in the recovery of blood flow perfusion and vessel density in ischemic muscles. However, a complete recovery of neovascularization was demonstrated in mice exposed to cigarette smoke that were treated with probucol or antioxidant vitamins. We found that antioxidant therapy in mice exposed to cigarette smoke was associated with a significant reduction of oxidative stress levels in the plasma and in ischemic muscles. In addition, endothelial progenitor cells (EPCs) exposed to cigarette smoke extracts in vitro showed a significant decrease in their angiogenic activities (migration, adhesion and homing into ischemic tissues) that was completely rescued by probucol and antioxidants vitamins. The goal of the second study was to investigate the potential role of NADPH oxidase (Nox2) in the modulation of ischemia-induced neovascularization in the context of aging. We found that the expression of Nox2 is increased by aging in ischemic muscles of control mice. This is associated with a significant reduction of blood flow recovery after ischemia in older compared to young control mice. We also demonstrated that the density of capillaries and arterioles is significantly reduced in ischemic muscles of older animals, whereas oxidative stress levels are increased. Nox-2 deficiency reduces oxidative stress levels in ischemic tissues and improves blood flow recovery and vascular densities in older animals. We also demonstrated that the functional activities of EPCs (migration and adhesion to mature endothelial cells) were significantly reduced in older compared to young mice. However, Nox2 deficiency is associated with preserved EPCs functional activities in older animals. We also demonstrated an age-dependent pathological increase of oxidative stress in EPCs that is not found in Nox2-deficient animals. The third study of the research project investigated the effect of fatty acids derived from the diet on postnatal neovascularization. To this end, mice received a diet containing either 20% corn oil (rich in omega-6) or 20% fish oil (rich in omega-3). Our results demonstrate that an omega-3 rich diet increases neovascularization in response to ischemia at the macrovascular, microvascular and clinical level compared to an omega-6 rich diet. This increased postnatal neovascularization is associated with decreased total cholesterol/HDL cholesterol ratio in the serum and improved VEGF/NO pathway in ischemic tissues. In addition, the omega-3 rich diet is associated with a significant increase of central (bone marrow) and peripheral (spleen) EPCs. We also show that the functional activities of EPCs (migration and incorporation into tubules) are improved and oxidative stress level in EPCs is reduced by the omega-3 rich diet. In conclusion, our studies have clarified the impact of cardiovascular risk factors (smoking and aging) and fatty acids derived from the diet (omega-3) on ischemia-induced neovascularization. We have also identified several mechanisms involved in that physiopathology. Globally, our studies should contribute to a better understanding of the effects of cigarette smoking, aging and omega-3 on the evolution of ischemic vascular diseases.

L’obésité est un facteur de risque lié à des problèmes physiques, émotionnels et comportementaux. Aujourd’hui, l’alimentation est composée d’un régime typiquement occidental «Western diet» qui est riche en acides gras saturés (AGS) et pauvre en acides gras polyinsaturés (AGPI) tel que les oméga-3 (N-3) et occasionnant un déséquilibre du ratio alimentaire N-6/N-3. Ce déséquilibre est une des causes de la prévalence des maladies mentales y compris celles des troubles de l'humeur et de l’anxiété. L’acide docosahexaénoïque (ADH, 22: 6 n-3) est l’acide gras (AG) le plus abondant dans le cerveau et son accumulation est particulièrement élevée pendant la période périnatale. Il joue un rôle important dans le développement neuronal et d'autres fonctions du cerveau tel l'apprentissage et la mémoire. Des perturbations de l’environnement périnatal peuvent influencer à très long terme l’avenir de la descendance en la rendant plus susceptible de développer des problèmes d’obésité dans un contexte nutritionnel riche. On ignore cependant si le déficit alimentaire chez la mère et particulièrement en ADH aura un impact sur la motivation alimentaire de la progéniture. L’objectif principal de cette thèse est d’étudier le rôle potentiel des N-3 sur la balance énergétique, la motivation alimentaire, la dépression et le niveau d’anxiété des descendants de souris mâles adultes assujetties à une alimentation riche en gras. Nos données ont démontré qu‘un régime maternel déficitaire en ADH durant la période périnatale incitait la descendance à fournir plus d’effort afin d’obtenir un aliment palatable. Ceci entraînerait un dérèglement de l’homéostasie énergétique en augmentant le gain de poids et en diminuant l’activité locomotrice tout en exacerbant le comportement de type anxieux dès que les souris sont exposées à un milieu obésogène. Les acides gras libres (AGL) sont des nutriments essentiels fonctionnant comme des molécules de signalisation dans le cerveau en ayant des récepteurs qui jouent un rôle important dans le contrôle du métabolisme énergétique. Parmi eux, on distingue un récepteur couplé à la protéine G (GPCR), le GPR120. Ce récepteur activé par les AGPI ω-3 intervient dans les mécanismes anti-inflammatoires et insulino-résistants via les N-3. Une mutation dans le gène GPR120 occasionnée par une réduction de l’activité de signalisation du gène est liée à l’obésité humaine. L'objectif premier de cette deuxième étude était d’évaluer l'impact de la stimulation pharmacologique de GPR120 dans le système nerveux central (SNC) sur l'alimentation, les dépenses d'énergie, le comportement de type anxieux et la récompense alimentaire. Nos résultats démontrent qu’une injection centrale aiguë d'agoniste GPR120 III réduit la prise alimentaire ad libitum et la motivation alimentaire pour un aliment riche en gras et en sucre; ainsi que les comportements de type anxieux. L’injection centrale chronique (21 jours) de ce même agoniste GPR120 III transmis par une pompe osmotique a démontré que les souris placées sous diète hypercalorique (HFD n’ont présenté aucune modification lors de la prise alimentaire ni de gain de poids mais qu’il y avait comparativement au groupe de véhicule, une réduction du comportement de type anxieux, que ce soit dans le labyrinthe en croix surélevé (LCS) ou dans le test à champ ouvert (OFT). L’ADH est reconnu pour ses propriétés anorexigènes au niveau central. De plus, la stimulation des récepteurs de GPR120 au niveau du cerveau avec un agoniste synthétique peut produire un effet intense intervenir sur le comportement lié à l'alimentation des rongeurs. Trouver une approche visant à contrôler à la fois la neuroinflammation, la récompense alimentaire et les troubles émotionnels aiderait assurément au traitement de l'obésité et du diabète de type 2. / Obesity is a risk factor for metabolic and mood disorders. The increasing abundance of the "Western diet" that is rich in saturated fatty acids (SFA) and low in polyunsaturated fatty acids (PUFA) omega-3 (N-3) can generate a physiological imbalance in the ratio of N-6/N-3 fatty acids. Such an imbalance has also been implicated in the increased prevalence of mood disorders and metabolic diseases. Docosahexaenoic acid (DHA, 22:6n-3), the most abundant fatty acid (FA) in the brain is accumulated not only during the post-natal period but also during the perinatal period. It plays an important role in neuronal development and other brain functions such as learning and memory. Disturbances of the perinatal environment can influence the sustainable future of the offspring, making it more likely to develop obesity in rich nutritional context. Some data suggest that an inadequate maternal intake of N-3 during pregnancy and the perinatal period may cause an increase in appetite signalling in the offspring as well as an increase rate of neurological and cardio-metabolic disease. It is not known if maternal dietary deficiency in N-3 will have an impact on food motivation of the offspring. The main objective of this thesis was to study the potential effect of dietary deficiency of DHA during the perinatal period on energy balance, food motivation and the anxiety-like behavior of adult male mouse offspring placed on HFD. Our data showed, as expected, that the maternal DHA deficient diet during perinatal period encouraged offspring to work harder to get a palatable food, entailed a dysregulation of energy homeostasis by increasing the body weight and reducing locomotor activity and exacerbated the anxiety-like behavior once they were exposed to an obesogenic environment. Free fatty acids (FFA) are essential nutrients that they also function as signalling molecules in the brain and have receptors that play a significant role in the control of energy metabolism. Among them is GPR120, also known as N-3 FA receptor, a g-protein coupled receptor that is reportedly activated by PUFA and shown to mediate the anti-inflammatory and insulin-sensitizing effects of N-3 FA. A mutation in the GPR120 gene that is associated with reduced GPR120 signalling activity is linked to human obesity. The objective of our second study was to test the impact of pharmacological GPR120 stimulation in the CNS on feeding, energy expenditure, anxiety-like behavior and food reward. Our results showed that an acute central injection GPR120 agonist III: reduced ad libitum food intake and the rewarding effect for high fat, high sugar food; produced a significant decrease in the anxiety-like behavior. While mice with the chronic injection (21 days) of GPR120 III agonist pump on HFD didn’t show any modification in food intake or body weight gain, but show a reduction of anxiety-like behavior in Elevated plus maze (EPM) and open-field test (OFT) compare to the vehicle group. DHA is notifies to have an anorectic action. Furthermore, stimulation of GPR120 receptors with synthetic agonist may cause an acute and profound effect on food related behavior in rodents. To find an approach that aims to control neuroinflammation, food reward and emotional disorders would greatly assist the treatment of obesity and type 2 diabetes.

Un faible apport alimentaire en acides gras polyinsaturés (AGPI) de la série n-3 a été associé à la prévalence des troubles de l'humeur chez l’Homme. Chez les rongeurs, les approches nutritionnelles visant à modéliser une alimentation pauvre en AGPI n-3 ont largement été développées au siècle dernier. En effet, un régime alimentaire carencé en AGPI n-3 sur une ou plusieurs générations induit chez le rongeur des altérations des comportements émotionnels tels que des comportements de type dépressif ou anxieux. Nous avons montré au laboratoire Nutrineuro que des souris nourries avec un régime déficient en AGPI n-3 présentent des niveaux d’AGPI n-3, en particulier l'acide docosahexaénoïque (DHA, un AGPI n-3) plus faible dans le cortex préfrontal (PFC) et dans le noyau accumbens (NAc) par rapport aux souris contrôle. De plus, nous avons pu mettre en évidence qu’une alimentation déficiente en AGPI n-3 est capable de moduler la plasticité synaptique dépendante du système endocannabinoïde (eCB). De fait, la réduction de DHA dans le CPF et le NAc est accompagnée d'une altération de la dépression à long terme (LTD-eCB) et des voies de signalisation dépendantes du système eCB au niveau du CPF (Lafourcade et al., 2011 ; Larrieu et al, 2012). Nos données indiquent que ces altérations sont dues à un découplage entre le récepteur cannabinoïde 1 (CB1R) et la protéine Gi/o. De plus, les souris déficientes en AGPI n-3 présentent des déficits comportementaux dans plusieurs tests évaluant les comportements émotionnels. Afin de mieux comprendre les mécanismes qui sous-tendent la diminution du DHA dans le CPF et les altérations des comportements émotionnels, nous avons étudié la morphologie neuronale dans le CPF et l’axe hypothalamo-hypophysaire (HPA) chez les souris déficientes en AGPI n-3. Nous avons montré que le régime alimentaire déficient en AGPI n-3 induit une atrophie de l’arborisation dendritique dans les neurones pyramidaux du CPF. L'atrophie dendritique est semblable à celle mesurée chez les souris soumises au régime équilibré en AGPI n-3 et soumises à un stress chronique de défaite sociale (CSDS). Aucun effet additionnel du CSDS sur la morphologie neuronale et le comportement émotionnel n’a été observé chez les souris déficientes en AGPI n-3. Nous avons ensuite étudié le rôle de l’axe HPA dans le développement des altérations comportementales et neurobiologiques chez les souris déficientes en AGPI n-3. Ces souris présentent une diminution de l'expression des récepteurs des glucocorticoïdes (GR) dans le CPF associée à une augmentation des taux circulants de corticostérone. Dans leur ensemble, nos résultats montrent qu’un faible apport alimentaire en AGPI n-3 peut modifier la plasticité synaptique dépendante du système eCB ainsi que l’arborisation dendritique des neurones du CPF. Nous avons également pu montrer que l’élévation des niveaux de corticostérone était impliquée dans l’altération des comportements émotionnels observée chez des souris nourries avec un régime déficient en AGPI n-3. / Low dietary intake of n-3 polyunsaturated fatty acids (PUFAs) has been associated with the prevalence of mood disorders in humans. In rodents, nutritional approaches aiming at modeling poor dietary n-3 PUFAs intake have been extensively developed in the last century. As a result, one- or multi-generation dietary n-3 deficiency induces depressive and anxiety-like behaviors. We have shown in the Nutrineuro lab that mice fed with a diet deficient in n-3 PUFAs exhibit decreased n-3 PUFAs levels, especially docosahexaenoic acid (DHA, a n-3 PUFA) levels in the prefrontal cortex (PFC) and in the nucleus accumbens (NAc). We showed that dietary n-3 PUFA is able to modulate endocannabinoid (eCB) dependent plasticity since DHA reduction in PFC and NAc is accompanied with eCB dependent long term depression (eCB-LTD) and eCB signaling impairment in the PFC (Lafourcade et al., 2011; Larrieu et al., 2012). Our data indicate that LTD alteration results from region-specific uncoupling of CB1 receptor from its effector Gi/o protein. In addition, n-3 deficient mice display behavioral deficits in several tests measuring emotional behavior. To further understand the mechanisms underlying DHA decrease in the PFC and emotional behavior alteration, we thoroughly investigated neuronal morphology and hypothalamic-pituitary-adrenal (HPA) axis in n-3 deficient mice. We showed that n-3 deficient diet induced dendritic atrophy in pyramidal neurons within the PFC. The dendritic atrophy was comparable to the one measured in control diet mice submitted to chronic social defeat stress (CSDS). No additional effect of CSDS on both neuronal morphology and emotional behavior was measured in n-3 deficient mice. We therefore investigated the role of the HPA axis deregulation in the development of behavioral and neurobiological alterations of n-3 deficient mice. We found a decreased expression of glucocorticoid receptor (GR) in the PFC of n-3 deficient mice together with increased circulating levels of corticosterone. Collectively, we unraveled one crucial mechanism underlying n-3 deficiency-induced alterations. Our results show that low dietary n-3 PUFAs can alter eCB-dependent plasticity and neuronal dendritic atrophy within the PFC leading to emotional behavior impairment. Importantly, we further demonstrated that corticosterone elevation in n-3 deficient mice was involved in the n-3 deficiency-induced emotional behavior and dendritic arborization alterations.

No presente estudo avaliamos o impacto da adição de fluido folicular (FF) de mulheres inférteis sem e com endometriose em estágios iniciais (I/II) e avançados [(III/IV) sem e com endometrioma] ao meio de maturação in vitro (MIV) sobre as taxas de normalidade meiótica de oócitos bovinos. Avaliamos se a L-carnitina (LC) e os ácidos graxos ômega-3 [n3, ácidos docosahexaenóico (DHA) e eicosapentaenoico (EPA)] são capazes de prevenir os danos meióticos em oócitos bovinos induzidos por FF de mulheres inférteis com endometriose I/II e III/IV durante a MIV. Para isso, realizamos um estudo experimental utilizando modelo bovino. Trinta e duas amostras de FF foram colhidas de 24 mulheres inférteis com endometriose (8 com I/II, 8 com III/IV sem endometrioma e 8 III/IV com endometrioma no ciclo) e 8 sem endometriose (controle) que foram submetidas à estimulação ovariana controlada para realização de injeção intracitoplasmática de espermatozoide. Complexos cumulus-oócitos (CCOs) imaturos de bovinos foram submetidos à MIV divididos em 9 grupos: sem FF (sem-FF), com 1% de FF de mulheres inférteis sem endometriose (FFControle) e com endometriose (FFEI/II, FFEIII/IV e FFEendometrioma) suplementados ou não com LC (0,6mg/mL) e ácidos graxos ômega-3 (0,4 nM de DHA e 0,6 nM de EPA) (FFControle+LC+n3, FFEI/II+LC+n3, FFEIII/IV+LC+n3 e FFEendometrioma+LC+n3). Após 22-24h de MIV, os oócitos foram denudados, fixados e armazenados para realização de imunofluorescência para visualização do fuso meiótico e cromossomos por microscopia confocal. As taxas de metáfase II (MII) e de MII normais foram comparadas entre os 9 grupos utilizando o teste do qui-quadrado (p<0,05). Um total de 1686 CCOs imaturos foram submetidos à MIV, e 1401 oócitos foram visualizados por microscopia confocal. A adição de FF de mulheres com endometriose ao meio de MIV reduziu a taxa de MII normais (FFEI/II: 62,2%, FFEIII/IV: 70,2% e FFEendometrioma: 72,7%) comparado aos grupos sem-FF (87,2%) e FFControle (87,2%). O grupo FFEendometrioma (69,3%) apresentou a menor taxa de MII comparado a todos os demais grupos (sem-FF: 91,9%, FFControle: 89,2%, FFControle+LC+n3: 89,2%, FFEI/II: 85,4%, FFEI/II+LC+n3: 85,3%, FFEIII/IV: 80,7%, FFEIII/IV+LC+n3: 90,8%, FFEndometrioma+LC+n3: 86,4%). O grupo FFEIII/IV apresentou menor taxa de MII comparado ao grupo sem-FF. No grupo com FFControle, a adição de LC+n3 não alterou as taxas de MII (89,2% vs 89,2) e de MII normais (87,2% vs 82,5%). No grupo FFEI/II, a adição de LC+n3 aumentou a taxa de MII normais (84,5% vs. 62,2%). No grupo FFEIII/IV a adição de LC+n3 aumentou a taxa de MII normais (70,2% vs 84,1%) e de MII (90,8%), que passou a ser semelhante a dos grupos sem-FF e FFControle. No grupo FFEendometrioma a adição de LC+n3 aumentou a taxa de MII normais (86,4%), comparado ao grupo FFEendometrioma (69,3%), a qual foi similar a dos grupos sem-FF e FFControle. Portanto, o FF de mulheres com endometriose prejudica o fuso meiótico e o alinhamento cromossômico de oócitos bovinos, independentemente, do estágio da doença. Entretanto, o avanço da endometriose e a presença de endometrioma parecem ter um impacto ainda mais negativo na qualidade oocitária, prejudicando também a maturação nuclear. A adição de LC+n3 previne os danos meióticos oocitários provocados pelo FF de mulheres com endometriose em estágios iniciais e avançados. Dessa forma, sugerimos que inflamação, o estresse oxidativo e a desregulação da ?-oxidação são fatores envolvidos na alteração da qualidade oocitária e, consequente, piora da fertilidade natural de mulheres com endometriose. / In the present study, we evaluated the impact of the addition of follicular fluid (FF) from infertile women without and with endometriosis in the early (I/II) and advanced stages [(III/IV) with and without endometrioma] to the in vitro maturation (IVM) medium on the meiotic normality rates of bovine oocytes. We evaluated whether L-carnitine (LC) and omega-3 fatty acids [n3, docosahexaenoic (DHA) and eicosapentaenoic acid (EPA)] were able to prevent bovine meiotic oocyte damage induced by FF from infertile women with endometriosis in stages I/II and III/IV during IVM. For this, we performed an experimental study using bovine model. Thirty-two FF samples were collected from 24 infertile women with endometriosis (8 with I/II, 8 with III/IV without endometrioma and 8 III/IV with endometrioma in the cycle) and 8 without endometriosis (control) who underwent to controlled ovarian stimulation for intracytoplasmic sperm injection. Immature cumulus oocytes complexes(COCs) of bovines were submitted to IVM divided into 9 groups: without FF (No-FF), with 1% FF of infertile women without endometriosis (FFControl) and with endometriosis (FFEI/II, FFEIII/IV and FFEendometrioma) supplemented or not with LC (0.6 mg/mL) and omega-3 fatty acids (0.4 nM DHA and 0.6 nM EPA) (FFControl+LC+n3, FFEI/II+LC+n3, FFEIII/IV+LC+n3 and FFEendometrioma+LC+n3). After 22-24h of IVM, the oocytes were denuded, fixed and stored for subsequent immunofluorescence to visualize the meiotic spindle and chromosomes by confocal microscopy. The metaphase II (MII) and normal MII rates were compared between the 9 groups using the chi-square test (p <0.05). A total of 1686 immature COCs were submitted to IVM, and 1401 oocytes were visualized by confocal microscopy. Addition of FF from women with endometriosis to the IVM medium decreased the rate of normal MII (FFEI/II: 62.2%, FFEIII/IV: 70.2% and FFEendometrioma: 72.7%) compared to the No-FF (87.2%) and FFControl (87.2%) groups. The FFEendometrioma group (69.3%) presented the lowest rate of MII compared to all other groups (No-FF: 91.9%, FFControl: 89.2%, FFControl+LC+n3: 89.2%, FFEI/II: 85.4%, FFEI/II+LC+n3: 85.3%, FFEIII/IV: 80.7%, FFEIII/IV+LC+n3: 90.8%, FFEndometrioma+LC+n3: 86.4%). The FFEIII/IV group had a lower MII rate compared to the No-FF group. In the group with FFControl, the addition of LC+n3 did not change the rates of MII (89.2% vs 89.2%) and normal MII (87.2% vs 82.5%). In the FFEI/II group, the addition of LC+n3 increased the normal MII rate (84.5% vs 62.2%). In the FFEIII/IV group, the addition of LC+n3 increased the normal MII rate (70.2% vs 84.1%) and MII (90.8%), which was similar to that of the No-FF and FFControl. In the FFEendometrioma group, the addition of LC+n3 increased the normal MII rate (69.3% vs 86.4%) which was similar to No-FF and FFControl groups. Therefore, the FF of women with endometriosis impairs the meiotic spindle and the chromosomal alignment of bovine oocytes, regardless of the stage of the disease. However, the progression of endometriosis and the presence of endometrioma appear to have an even more negative impact on oocyte quality, and also impairs nuclear maturation. The addition of LC+n3 prevents meiotic oocyte damages induced by FF from women with endometriosis in the early and advanced stages. Thus, we suggest that inflammation, oxidative stress and deregulation of ?-oxidation are factors involved in the alteration of oocyte quality and, consequently, worsening of the natural fertility of women with endometriosis.

Introdução: A terapia antirretroviral (ART) mudou o curso da Aids, porém está associada a alterações metabólicas e aumento do risco de doenças cardiovasculares. Objetivo: Avaliar o efeito da suplementação com ácidos graxos ômega-3 de origem marinha no perfil lipídico, na homeostase da glicose, na distribuição de gordura corporal e nos marcadores inflamatórios de adultos com HIV em ART. Métodos: Artigo 1. Trata-se de uma revisão sistemática da literatura com metanálise. Realizou-se busca por ensaios clínicos na base de dados PubMed; 33 artigos foram localizados, seis cumpriram os critérios de inclusão e quatro apresentavam qualidade metodológica adequada. Foi realizada metanálise com efeitos fixos e descrição das diferenças de médias sumárias (DMS (IC95 por cento )). Artigos 2 e 3. Trata-se de um ensaio clínico aleatorizado e controlado. Foram recrutados 120 adultos com idade entre 19 e 64 anos, de ambos os sexos. Os indivíduos alocados no grupo intervenção foram suplementados por 24 semanas com 3g de óleo de peixe/dia (900mg de ácidos graxos ômega-3) e indivíduos alocados no grupo controle receberam placebo (óleo de soja). Resultados: Artigo 1. Após 8-16 semanas de intervenção com 900-3360mg de ácidos graxos ômega-3/dia, observou-se redução de -80,34mg/dL (IC 95 por cento : -129,08 a -31,60) nas concentrações de triglicérides. A análise agregada de estudos com média de concentração de triglicérides > 300mg/dL no baseline e intervenção com 1800-2900mg de ácidos graxos ômega-3/dia resultou em redução de -129,72mg/dL (IC95 por cento : -206,54 a -52,91). Artigos 2 e 3. Foram considerados nas análises dados de 83 sujeitos. Os modelos multinível não revelaram relação estatisticamente significante entre a suplementação com óleo de peixe e as mudanças longitudinais nas concentrações de triglicérides (p=0,335), LDL-C (p= 0,078), HDL-C (p=0,383), colesterol total (p=0,072), apo B (p=0,522), apo A1 (p=0,420), razão LDL-C/apo B (p=0,107), índice homa-2 IR (p=0,387), IMC (p=0,068), circunferência da cintura (p=0,128), relação cintura/quadril (p=0,359), PCR ultra sensível (p=0,918), fibrinogênio (p=0,148), e fator VIII (p=0,073). Conclusões: Artigo 1. Diferentes doses de ácidos graxos ômega-3 reduziram modo significativo as concentrações de triglicérides, confirmando a potencial aplicabilidade desse nutriente no tratamento da hipertrigliceridemia em pessoas que vivem com HIV em ART. Artigos 2 e 3. Uma dose relativamente baixa de óleo de peixe para pessoas que vivem com HIV em ART não alterou o perfil lipídico, a homeostase da glicose, a distribuição de gordura corporal e a concentração de marcadores inflamatórios. Recomenda-se, em estudos subseqüentes, a avaliação do efeito de doses mais elevadas, bem como a determinação de marcadores inflamatórios mais sensíveis / Background: Although the antiretroviral therapy (ART) revolutionized the care of HIV-infected subjects, it has been associated with metabolic abnormalities and increased risk of cardiovascular diseases. Aims: To review the effects of marine omega-3 fatty acids on lipid profile, insulin resistance and inflammatory markers in subjects living with HIV on ART. Methods: Paper 1. Thirty three articles were found in a PubMed search; six met the inclusion criteria; and four of them were considered of adequate quality and included. Meta-analysis with fixed effects was performed and weighted mean differences (WMD (95 per cent CI)) were described. Paper 2 and 3. The study was conducted in an HIV/Aids care centre affiliated to the Medical School, University of Sao Paulo. This was a randomized controlled trial that assessed the effects of 3g fish oil/day (900mg of omega-3 fatty acids) or 3g soy oil/day (placebo). A hundred and twenty subjects aged between 19 and 64 years were recruited. The statistical analyses were performed in Stata 9. Results: Paper 1. Data from 83 subjects were included in the analyses. The overall reduction on triglyceride concentrations after 8-16 weeks of treatment with 900-3360mg of omega-3/day was WMD=-80.34mg/dL (95 per cent CI: -129.08 to -31.60). The pooled result of studies with mean triglyceride > 300 mg/dL at baseline and 1800-2900mg omega-3/day was WMD=-129.72mg (95 per cent CI: -206.54 to -52.91). Paper 2 and 3. Multilevel analyses revealed no statistically significant relationships between fish oil supplementation and the longitudinal changes in triglyceride (p= 0.335), LDL-C (p= 0.078), HDL-C (p= 0.383), total cholesterol (p=0.072), apo B (p= 0.522), apo A1 (p=0.420), LDL-C/apo B ratio (p=0.107), homa-2 IR index (p=0.387), BMI (p=0.068), waist circumference (p=0.128), waist/hip ratio (p=0.359), hs-CRP (p=0.918), fibrinogen (p=0.148), and VIII factor (p=0.073). Conclusions: Paper 1. Different doses of omega-3 fatty acids reduced significantly triglyceride concentrations confirming the potential applicability of this nutrient on the management of hypertriglyceridemia in HIV-infected subjects on ART. Paper 2 and 3. A relatively low dose of fish oil for HIV subjects on ART did not change lipid profile, insulin resistance, body fat distribution, and inflammatory markers. Further investigations should considerer the assessment of higher doses and more sensitivity inflammatory markers

Introdução: Existem poucas estratégias de intervenção medicamentosa que se mostraram eficazes na doença hepática gordurosa não alcoólica (DHGNA). Os ácidos graxos poliinsaturados (AGPI) Ômega-3 parecem ser eficazes no tratamento da esteatose hepática de modelos experimentais, mas poucos estudos randomizados em humanos foram realizados. O objetivo deste estudo foi avaliar prospectivamente a eficácia de AGPI Ômega-3 provenientes do óleo de linhaça e peixe na esteatohepatite não alcoólica (pacientes com EHNA). Métodos: Sessenta pacientes com biópsia confirmando EHNA foram incluídos no estudo duplo cego, randomizado, placebo controlado. Os pacientes foram randomizados em dois grupos. Grupo Ômega-3 (n = 32) recebeu três cápsulas contendo no total 945 mg de AGPI Ômega-3 (63% ácido alfa linolênico (ALA), 21% ácido eicosapentaenóico (EPA) e 16% do ácido docosahexaenóico (DHA)) e Grupo Placebo (n = 28) recebeu três cápsulas contendo óleo mineral. A intervenção foi realizada por seis meses, quando os pacientes foram novamente submetidos à biópsia hepática. O desfecho primário foi a mudança histológica hepática de acordo com o escore de atividade EHNA (NAS) no início (pré intervenção) e seis meses (após intervenção). Desfecho secundário compreendeu análise das aminotransferases séricas, perfil lipídico e glicemia em jejum, parâmetros antropométricos e nível sérico de IL6 em 0, 3 e 6 meses e dosagem de Ômega-3 plasmático, como prova de tratamento, em 0 e 6 meses. Resultados: Dos 60 pacientes avaliados, 10 não terminaram o estudo (5 no grupo Ômega-3, e 5 no grupo placebo). Analisando os resultados primários, a atividade NAS melhorou ou se manteve estável em 78,26% dos pacientes do grupo placebo e em 55,56% do grupo Ômega-3, sem diferença significativa entre os grupos (p = 0,978), a inflamação lobular reduziu ou se manteve estável em 91,3% no grupo de placebo e em 66,67% no grupo Ômega-3, também sem diferença entre os grupos (p = 0,994). Ômega-3 não reduziu a esteatose hepática, balonização hepatocelular e fibrose. Nos parâmetros bioquímicos houve redução do nível de triglicérides em três meses no grupo Ômega-3 (p = 0,011) quando comparado com o placebo. Por outro lado, aminotransferases séricas, colesterol total e frações, glicemia, parâmetros antropométricos e níveis séricos de IL-6 não foram alterados com o tratamento em seis meses quando comparado com placebo. Analisando o resultado do Ômega-3 plasmático no período seis meses, observou-se que no grupo Ômega-3 houve aumento do ALA (p = 0,014), EPA (p = 0,016), da relação Ômega-3/Ômega-6 (p = 0,018) e redução do ARA (p= 0,028), enquanto que no grupo placebo encontrou-se aumento do Ômega-3 nas formas de EPA (p = 0,03), DHA (p = 0,036) relação Ômega-3/Ômega-6 (p = 0,007), o que comprova que o grupo placebo de alguma forma ingeriu também Ômega-3. Avaliando o Ômega-3 plasmático entre grupos tratamento e placebo tem-se diferença entre os grupos em relação ao ARA (p = 0,03) que reduziu no grupo \"tratado com Ômega-3\". Devido ao consumo de Ômega-3 pelo grupo placebo, optou-se por desconsiderar o duplo cego e fazer nova análise estatística baseada no aumento de Ômega-3 plasmático e comparar com melhora histológica das variáveis NAS e encontrou-se que o aumento do DHA estava positivamente relacionado com a melhora ou estabilização da inflamação lobular em seis meses de estudo (p = 0,014). Conclusões: Os resultados dste estudo indicam que AGPI Ômega-3 a partir de uma mistura de óleos de linhaça e peixe não pode melhorar a histologia hepática, a maioria dos parâmetros bioquímicos e os níveis séricos de IL-6; no entanto, esta suplementação impacta significativamente o perfil lipídico dos pacientes com EHNA, aumentando os níveis de AGPI Ômega-3, diminuindo os níveis da ácido araquidônico (AA), potencialmente próinflamatória da família AGPI Ômega-6, e diminuição dos níveis de triglicérides séricos. Na análise post hoc houve correlação significativa entre o aumento dos níveis plasmáticos de DHA e melhora da inflamação lobular, independentemente do tratamento recebido. A limitação do estudo foi o grupo placebo ter ingerido ácidos graxos Ômega-3. Mais estudos são necessários para confirmar estes resultados (ID 01992809) / Introduction: There is a limited number of effective drug treatments available for the treatment of nonalcoholic steatohepatitis (NASH). Polyunsaturaturated fatty acid (PUFAs) Omega 3 seems to be effective in treating hepatic steatosis in experimental animal models, however there is a limited number of humans randomized studies available in the literature. The purpose of the present study is to prospectively evaluate the treatment efficacy of the PUFAs Omega 3 from flaxseed and fish in patients with NASH disease. Methods: A total of sixty biopsy confirmed NASH patients were included in a double blind, randomized, placebo controlled study. They were randomized into two groups: Omega 3 group (n = 32) where patients received a total of 945mg of PUFAs Omega 3 and Placebo group (n = 28) where patients received only mineral oil. After a 6 month treatment all patients underwent a new liver biopsy. Primary goal was to evaluate and compare liver histologic changes, according to Nonalcoholic fatty liver disease (NAFLD) activity score (NAS), between pre and post treatment biopsies. Secondary goal was to evaluate serum transaminases, lipid profile and serum non fasting glucose, anthropometric parameters and serum IL6 at 0, 3 and 6 month treatment. A serum Omega 3 dosage was performed at 0 and 6 month as treatment proof. Results: A total of 50 patients finished the study, 25 from each original group. NAS score improved or was unaltered in 78.26% of the placebo group and in 55.56% of the Omega 3 group (p = 0,978). Lobular liver inflammation was reduced or unaltered in 91.3% and in 66.67% respectively of the placebo and Omega 3 groups (p = 0,994). Omega 3 alone was not able to reduce liver steatosis, hepatocelular balonization or fibrosis. Biochemical analysis revealed reduction on serum triglycerides after 3 month treatment for the Omega 3 group patients, when compared to placebo (p=0,011). Serum transaminases, total cholesterol and fractions, non fasting glucose and IL-6 were found to have no changes after 6 month treatment, when compared to placebo. After 6 month we found serum rise of Omega 3 on its forms as ALA (p= 0,014) and EPA (p = 0,016), and of the relation Omega 3/Omega 6 (p = 0,018), besides a reduction of the ARA (p= 0,028). The placebo group demonstrated to have an Omega 3 serum rise of its forms as EPA (p = 0,03) and DHA (p = 0,036) and of the relation Omega3/Omega6 (p = 0,007). These findings are proof that the placebo group also ingested some form of Omega 3. There is a difference between the groups regarding the serum Omega 3 on the ARA relation, which was reduced in the Omega 3 group. Due to the Omega 3 ingestion by the placebo group we decided to not consider the double blind and to perform a new statistic analysis based on the serum Omega 3 rise and compare with the improvement on NAS histologic variables. The analysis revealed that DHA rise was positively related with an improvement or unchanged of lobular inflammation after 6 months (p= 0,014). Conclusions: The present study was able to demonstrate that the AGPI Omega 3 from flaxseed and fish can not improve hepatic histology, most of the biochemical parameters and serum levels of IL-6. However this type of supplementation revealed a significant impact over lipid profile from NASH patients, providing an rise on AGPI Omega 3 levels and reducing the levels of Araquidonic Acid (AA) and triglycerides. The post hoc analysis demonstrated a significative correlation between the serum rise of DHA and the improvement of lobular inflammation, regardless of the received treatment. The fact that the placebo group ingested Omega 3 revealed to be a limitation of the present study. More studies are recommended to confirm our findings (ID 01992809)