Dissertations / Theses on the topic 'Nitric oxide resistance'

The insulin resistance syndrome [INSR] is associated with increased cardiovascular risk and affects up to 25% of the Australian population. The mechanism underlying the relationship between the INSR and increased cardiovascular risk is controversial. We postulated that perturbations in the renin-angiotensin system [RAS] and endothelium-derived NO may be implicated in the development of early vascular changes in the INSR. Repeated measurements of arterial stiffness [using digital photoplethysmography] and haemodynamic parameters in response to vasoactive medications were used to demonstrate the functional expression of angiotensin II [Ang II] receptors and NO synthase [NOS]. Ang II acts via two main receptor sub-types: the Ang II type 1 [AT1] and Ang II type 2 [AT2] receptors. The AT1 receptor is central to the development of arterial stiffness and endothelial dysfunction. The role of AT2 receptors in humans is controversial but is postulated to counter-act AT1 receptor mediated effects in diseased vascular beds. We demonstrated increased AT1 and AT2 receptor-mediated effects in small to medium-sized arteries of subjects with early INSR [Chapter 6]. In addition, functional expression of AT2 receptors in adult insulin resistant humans [Chapter 5], but not in healthy volunteers [Chapter 4] was demonstrated. AT1 receptor blockade in subjects with early INSR resulted in improvements in vascular function, with a consequent functional down-regulation of AT2 receptors [Chapter 7]. Functional NOS expression was demonstrated to be increased in subjects with early INSR compared with healthy controls [Chapter 6]. This was postulated to be a homeostatic response to counteract early vascular changes in subjects with early INSR. AT1 receptor blockade in these subjects reduced functional NOS expression [Chapter 8]. In conclusion, patients with early INSR represent a model of early disease where early intervention may be able to reverse the process incited by the initial exposure to multiple cardiovascular risk factors. Early vascular changes in these individuals are mediated at least in part, by increased AT1 receptor activity and/or expression, and may be detected by changes in arterial stiffness indices and non-invasive vascular reactivity studies. There is a compensatory increase in AT2 receptor and NOS expression/activity to counter-act these vascular changes.

Coronary blood flow (CBF) is principally controlled by changes in coronary vascular resistance (CVR). Low CVR helps to maintain myocardial perfusion in the presence of epicardial stenosis, therefore factors that impair the reduction of CVR will have a direct effect on CBF leading to either a narrower “effective” perfusion pressure range or reduced ability to compensate for increased demand on myocardial contraction. There are several mechanisms which may be important in the control CVR in humans including both endothelial dependent production of NO and reduction of the simple anion inorganic nitrite (a metabolite of NO) back to NO (via several putative mechanisms). The synthesis of NO by both nitric oxide synthase (NOS) and the reduction of nitrite in the coronary circulation has been the subject of many animal and human clinical studies. Reduced systemic endothelial dependent production of NO has an association with worse cardiovascular outcomes in humans, the number of potential mechanisms are large and perhaps central is the effect on CVR. The reduction of nitrite to NO is seen, in some ways, to be the perfect compensatory mechanism, particularly when endothelial function is impaired. It is easy to hypothesise that this stoichiometric balance of NO production might be responsible for the perfect regulation of CVR and thus CBF. Methods This thesis investigates the influence and effect of both endothelial production of NO and the reduction of nitrite to NO on CVR in man. First, an observational study assessing the impact of these mechanisms in patients undergoing percutaneous coronary intervention (PCI), in the treatment of both non-ST-elevation myocardial infarction and stable angina. Specifically, the metabolites of NO were measured from aortic root to coronary sinus together with the associated CVR both before and after PCI. Second, using a systemic infusion of sodium nitrite (NaNO2) in NSTEMI patients to assess the effect of nitrite reduction on CVR during PCI. The systemic nitrite concentration was increased 8-fold in the same experimental conditions as the observational study. Third, beyond CVR control the influence of NO and nitrite was also assessed in terms of platelet reactivity and systemic inflammatory cytokines in the NSTEMI cohort both with and without NaNO2 infusion. Results NSTEMI patients have a net increase in NO metabolites across the coronary circulation unlike healthy controls (historical data) and stable angina patients. This net increase is lost following PCI and is associated with a significant rise in CVR. Stable angina patients appear to compensate with increase collateral circulation and not NO synthesis. An 8-fold increase in nitrite concentration has no effect on CVR or platelet reactivity in NSTEMI patients. Conclusions In NSTEMI patients a net aorta to coronary sinus NO synthesis appears to be important to maintain a low CVR and thus CBF when haemodynamically significant epicardial disease is present. After the epicardial disease is treated this net increase in NOx (Nitric Oxide metabolites), is lost and is associated with an acute increase in CVR. Stable angina patients have no net increase in NOx across the coronary circulation and after revascularisation have no change in CVR, this may reflect an alternate mechanism of compensation and microvascular perfusion maintained by collateral circulation as evident by the increase CFI. Despite the perfect environment for the reduction of nitrite to NO we saw no influence of an 8-fold increase in serum nitrite concentration on CVR in patients with NSTEMI either before or after PCI, suggesting that nitrite reduction to NO plays no role in CBF regulation in NSTEMI patients. Nitrite reduction depends on conditions that are found predominantly in the capillary bed or venules, thus any mechanism would need to rely on a feedback mechanism to signal back to the arterioles (where much of resistance change is created). Despite hypotheses by others that this may occur by the close arrangement of venules to arterioles, this appears not to be the case in NSTEMI patients. Other clinically relevant and important mechanisms also appear to be unaffected by this increase in serum nitrite, residual platelet function and cytokine concentrations at 24 hours.

Orientador: Mario Jose Abdalla Saad
Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas
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Resumo: A resistência seletiva à insulina através da via IRS/PI3-K/Akt/eNOS associada à ativação normal ou exacerbada da via de crescimento MAPK tem sido proposta como um possível elo entre situações de resistência à insulina e doença cardiovascular. Inicialmente demonstramos que animais com resistência à insulina e doença cardiovascular (o rato espontaneamente hipertenso, SHR) apresentam menor ativação da via IRS/PI3-K/Akt/eNOS e hiperativação/hiperexpressão da via da MAPK na aorta torácica, enquanto a ativação normal da via IRS/PI3-K/Akt/eNOS pode proteger o animal obeso, resistente à insulina e que não apresenta doença cardiovascular. Posteriormente, outras vias estimulatórias do crescimento celular, como a via JAK/STAT, foram estudadas no vaso de outro modelo animal de resistência à insulina e doença cardiovascular - o rato com obesidade induzida por dieta. Este modelo demonstrou que a redução da ativação da via PI3-K/Akt/eNOS ocorre em paralelo à hiperativação das vias da MAPK e JAK/STAT, e isso pode desempenhar função importante da patogênese da doença cardiovascular neste estado patológico. Em outro estudo foi demonstrado pela primeira vez que a acetilcolina pode ativar a eNOS de maneira cálcio-independente, através da via IRS/PI3-K/Akt utilizando para isso uma tirosina quinase intracelular, a JAK2. Em ratos com obesidade induzida por dieta, resistentes à insulina e com disfunção endotelial, foi demonstrado que há resistência na via da PI3-K/Akt/eNOS tanto em resposta à insulina quanto à acetilcolina, em decorrência de maior fosforilação inibitória do IRS-1 e da redução dos níveis teciduais da eNOS. Assim, o desequilíbrio entre a ativação reduzida da via IRS/PI3-K/Akt/eNOS e a maior ativação das vias de crescimento (MAPK e JAK/STAT) pode contribuir para o desenvolvimento de doença cardiovascular em estados de resistência à insulina
Abstract: The actions of acetylcholine on endothelium are mainly mediated through muscarinic receptors, which are members of the G protein-coupled receptor family. In the present study we show that acetylcholine induces rapid tyrosine phosphorylation and activation of Janus kinase 2 (JAK2) in rat aorta. Upon JAK2 activation, tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1) is detected. In addition, acetylcholine induces JAK2/IRS-1 and IRS-1/phosphatidylinositol (PI) 3-kinase associations, downstream activation of Akt/protein kinase B, endothelial cell-nitric oxide synthase (eNOS), and extracellular signal-regulated kinase (ERK1/2). The pharmacological blockade of JAK2 or PI 3-kinase reduced acetylcholine-stimulated eNOS phosphorylation, NOS activity and aorta relaxation. These data indicate a new signal transduction pathway for IRS-1/PI 3- kinase/Akt/eNOS activation and ERK1/2 by means of JAK2 tyrosine phosphorylation stimulated by acetylcholine in vessels. Moreover, we demonstrate that, in aorta of obese rats (high-fat diet), there is an impairment in insulin- and acetylcholinestimulated IRS-1/PI 3-kinase pathway, leading to reduced activation with lower protein levels of eNOS associated with a hyperactivated ERK/MAP kinase pathway. These results suggest that in aorta of obese rats, there is not only insulin resistance, but also acetylcholine resistance, probably mediated by a common signaling pathway that controls the activity and the protein levels of eNOS
Doutorado
Medicina Experimental
Doutor em Fisiopatologia Medica

Orientador: Edson Antunes
Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas
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Resumo: A prevalência de obesidade vem aumentando a cada ano, sendo um dos principais problemas de saúde pública. A obesidade é considerada importante fator de risco para o surgimento de resistência à insulina, dislipidemia, diabetes tipo 2 e hipertensão arterial. Estes fatores podem atuar de maneira sinérgica no desenvolvimento da disfunção erétil. Entretanto, até o momento, poucos estudos procuraram avaliar os mecanismos envolvidos na disfunção erétil causada pela obesidade. Por isso, utilizando o modelo de obesidade induzida por dieta hiperlipídica em camundongos, o presente trabalho procurou caracterizar a disfunção erétil nestes animais. Para tanto, utilizamos experimentos funcionais in vivo e in vitro, assim como ensaios bioquímicos e morfológicos. Especificamente, realizamos os seguintes experimentos nos corpos cavernosos dos camundongos controles e tratados com dieta hiperlipídica por 10 semanas: 1) curvas concentração-efeito in vitro a agentes relaxantes dependente (acetilcolina) e doador de óxido nítrico (nitroprussiato de sódio), assim como à estimulação elétrica (relaxamento nitrérgico); 2) curvas concentração-efeito in vitro a agentes contráteis como fenilefrina (agonista 'alfa' 1-adrenérgico) e endotelina-1, assim como a estimulação elétrica (contração neurogênica); 3) medida da pressão intracavernosa in vivo; 4) determinação dos níveis de GMPc; e 5) análise histomorfométrica. Os animais tratados com dieta hiperlipídica apresentaram aumento do peso corporal, da gordura epididimal, dos níveis séricos de colesterol total e LDL, e resistência à insulina. Os animais tratados com dieta hiperlipídica apresentaram redução da pressão intracavernosa (in vivo). Os relaxamentos nitrérgico e dependente de endotélio foram menores nos corpos cavernosos dos camundongos obesos. Por outro lado, as respostas contrateis induzidas pela fenilefrina, endotelina-1 e estimulação elétrica foram maiores nestes animais. Os níveis de GMPc nos corpos cavernosos dos animais tratados com dieta hiperlipídica foram significantemente menores do que nos tecidos de animais controles. Os corpos cavernosos dos animais tratados com dieta hiperlipídica não sofreram alterações histomorfométricas. Para investigar o efeito da resistência à insulina na função erétil, tratamos os animais com o sensibilizador de insulina, metformina, por 14 dias (300 mg/Kg/dia). O tratamento com metformina restaurou o relaxamento nitrérgico, o relaxa- mento dependente de endotélio, e o aumento da resposta contrátil à fenilefrina dos corpos cavernosos dos animais tratados com dieta hiperlipídica, sem alterar o peso corporal e a gordura epididimal. Em suma, nossos resultados mostram que o tratamento com dieta hiperlipídica por 10 semanas causa redução da pressão intracavernosa, assim como do relaxamento nitrérgico e dependente de endotélio e, concomitantemente, aumento da resposta contrátil in vitro. O tratamento com metformina reverteu a sensibilidade à insulina e o quadro de disfunção erétil, sugerindo que a disfunção erétil nos camundongos tratados com dieta hiperlipídica é resultante da resistência à insulina associada à obesidade
Abstract: The prevalence of obesity is increasing every year, and a becoming a major public health problem. Obesity is considered an important risk factor for development of insulin resistance, dyslipidemia, diabetes mellitus type 2 and arterial hypertension. These are risk factors that act synergistically in the development of erectile dysfunction. However, so far few studies have investigated the mechanisms involved in erectile dysfunction caused by obesity. Therefore, using a model of obesity induced high-fat diet in mice, the present study searched to characterize erectile dysfunction in these animals. Thus, we used functional experiments in vivo and in vitro, biochemical and morphological test. Specifically, the following experiments were performed in corpus cavernosum of control and high-fat fed mice. 1) concentration response curves in vitro to relaxing agents endothelium-dependent (acetylcholine) and NO donor (sodium nitroprusside), as well as to electrical-field stimulation (EFS) (nitrergic relaxations); 2) Concentration-response curves in vitro to the contractile agents phenylephrine ('alfa'-1 adrenergic receptor agonist) and endothelin-1, as well as to EFS (neurogenic contraction; 3) measurement of intracavernous pressure in vivo; 4) determination of cGMP levels; 5) hystomorfometric measurement. Mice treated with high-fat diet showed increased body weight, epididimal fat, total choles-terol and LDL, and insulin resistance. The corpus cavernosum of high-fat fed mice showed impaired intracavernous pressure. The endothelial and nitrergic relaxations were lower in corpus cavernosum of high-fat fed mice. Moreover, the contractile responses induced by phenylephrine, endothelin-1 and electrical stimulation were increase in this animals. The cGMP levels were lower in corpus cavernosum of high-fat fed mice in comparison with control mice. There were no morphological alterations in erectile tissue of mice treated with high-fat diet. To investigate the effect of insulin resistance in erectile function, we treated the animals with insulin sensitizer, metformin, for 14 days (300 mg /kg/day). Metformin treatment restored the impaired endothelium-dependent and nitrergic relaxations, and the enhanced contractile response to phenylephrine in the corpus cavernosum of high-fat fed mice, without affecting the body weight and epididymal fat. In conclusion, our data shows that treatment with high-fat diet for 10 weeks reduces the intracavernous pressure, as well as the endothelial and nitrergic cavernosal relaxations. Concomitantly, this treatment increases the contractile responses, in vitro. Metformin treatment restored the insulin sensitivity and the erectile dysfunction, suggesting that erectile dysfunction in high-fat fed mice is the result of insulin resistance associated with obesity.
Mestrado
Farmacologia
Mestre em Farmacologia

Thesis (MScMedSc (Biomedical Sciences. Medical Physiology))--University of Stellenbosch, 2009.
Insulin stimulates the production of nitric oxide (NO) in endothelial cells and cardiac myocytes by a signalling pathway that involves the insulin receptor substrate (IRS)-1, phosphatidylinositol-3-kinase and protein kinase B (PKB/Akt). Physiological concentrations of NO play an important part in maintaining normal vascular function. It has been suggested that nitric oxide synthase (NOS) activity and NO production are chronically impaired in diabetes mellitus by an unknown mechanism. The reninangiotensin system and subsequent production of angiotensin II (Ang II) are elevated in obesity and diabetes while antagonism of the AT1 receptor with Losartan has beneficial effects in patients with insulin resistance and type II diabetes. Aims: We therefore aimed to investigate (i) the effect of Ang II on myocardial insulin signalling with regards to key proteins (IRS-1, PKB/Akt, eNOS and p38 MAPK) in correlation with NO production, (ii) the effect of Losartan on these parameters. Methods: Hyperphagia-induced obese, insulin resistant rats (DIO=diet supplemented with sucrose and condensed milk) were compared to age-matched controls. Half the animals were treated with 10mg/kg Losartan per day for 1 week. Isolated hearts were perfused with or without 0.03 μIU/mL insulin for 15 min. Blood glucose, bodyweight, intraperitoneal fat and plasma insulin and Ang II were recorded. Proteins of interest and their phosphorylation were determined by Western blotting. NO production was flow cytometrically analyzed. ANOVA followed by the Bonferroni correction was used with a p< 0.05 considered significant. Results: DIO animals had significant elevated bodyweight, blood glucose, plasma insulin and Ang II levels. Our data showed that the hearts from the DIO animals are insulin resistant, ultimately reflected by the attenuated activation of the key proteins (IRS-1, PKB/Akt and eNOS) involved in insulin signalling as well as NO production. AT1 receptor antagonism improved NO production in isolated adult ventricular myocytes from DIO animals while concurrently enhancing expression of eNOS, PKB/Akt and p38 MAPK. In contrast, NO production as well as expression of eNOS and PKB/Akt was attenuated in control animals after Losartan treatment. Conclusion: These results suggested that Ang II via AT1 or AT2 receptors, modulates protein expression of both PKB/Akt and eNOS. This encouraged us to investigate the involvement of AT2 receptors in the observed changes. To investigate this we needed to establish a culture of neonatal rat cardiac myocytes treated with raised fatty acids and Ang II. If similar changes were induced as observed in the hearts of DIO animals, the involvement of the AT1 and AT2 receptors could be investigated using specific antagonists against these receptors. Primary cultured ventricular myocytes were isolated from 1-3 day old Wistar rat pups. They were cultured for 48 hours before the addition of palmitate and oleate at a concentration of 0.25 mM each and were treated with or without the fatty acids for a period of 4 days. After 18 hours of serum starvation, cells were stimulated with or without 10 nM insulin for 15 minutes. The effect of fatty acid treatment on cell viability and glucose uptake were assessed by trypan blue and propidium iodide staining and 2-deoxy-D-3[H] glucose uptake respectively. Protein levels and phosphorylation of key proteins (PKB/Akt, PTEN and p38 MAPK) in insulin signalling was determined by Western blotting. 0.25 mM Fatty acids did not result in the loss of cell viability. Contrary to expectation, fatty acid treatment led to enhanced basal glucose uptake but lower Glut 1 protein expression. Basal protein expression of PPARα was, however, upregulated as was the expression of the phosphatase, PTEN. The latter could explain the lower PKB/Akt phosphorylation also documented. From these results we conclude that neonatal cardiac myocytes, cultured in the presence of elevated fatty acids, did not respond in a similar manner as the intact hearts of our animals and further modifications of the system might be needed before it can be utilized as initially planned.

Le monoxyde d'azote NO• est impliqué dans de nombreux processus biologiques. Il intervient, entre autres, dans la vasodilatation, la neurotransmission, il peut impliquer le développement ou l'apoptose des cellules et possède également des propriétés bactéricides. Le contrôle de la libération de ce radical est donc de grand intérêt pour des applications biomédicales en chimiothérapie photo-activée (PACT) ainsi qu'en inactivation photo-dynamique (PDI). La stratégie ici est de synthétiser des complexes de ruthénium à ligand nitrosyle photoréactifs, qui sont capables de libérer NO• sous irradiation mono ou biphotonique. L'excitation à deux photons permet une irradiation dans la fenêtre thérapeutique, très focalisée et une pénétration du faisceau plus profonde qu'en monophotonique. Ces travaux de thèse sont consacrés à la synthèse et l'étude des propriétés photochimiques de complexes [RuNO] et à leurs applications en biologie. Le premier chapitre de cette thèse développe l'état de l'art dans le domaine des complexes de ruthénium à ligand nitrosyle et présente les enjeux biologiques. Le second chapitre présente les propriétés de photolibération de NO• de complexes possédant le ligand 4'-(2-fluorényl)-2,2':6',2''-terpyridine, sous excitation à un et à deux photons par des études spectroscopiques. Les photoproduits obtenus sont caractérisés par diffraction des rayons X. Dans un troisième chapitre, l'étude des complexes cis (Cl,Cl)- et trans (Cl,Cl)-[RuII(fluorène-terpyridine)Cl2NO]PF6 est menée dans l'eau. Les capacités de photolibération du trans (NO,OH)-[RuII(fluorène-terpyridine)(Cl)(OH)(NO)]PF6 dans les conditions biologiques sont étudiées. Le quatrième chapitre s'intéresse à la synthèse de nouveaux complexes constitués de ligands dérivés du 4'-(2-fluorényl)-2,2':6',2''-terpyridine et à leurs propriétés de photolibération de NO•. Le cinquième chapitre s'intéresse aux propriétés phototoxiques de ces complexes envers des cellules cancéreuses (HCT 116 et FaDu). Enfin, dans le sixième chapitre, les propriétés remarquables de ces systèmes dans la levée de la résistance de Staphylococcus epidermidis aux antibiotiques sont exposées
Nitric oxide NO• is involved in numerous biological processes. It takes part to vasodilatation, neurotransmission, it can trigger cell proliferation or apoptosis and it also has bactericidal properties. Thus, NO• release control is of high interest for biomedical applications such as photo-activated chemotherapy (PACT) or photodynamic inactivation (PDI). The strategy here is to synthesize photoreactive ruthenium complexes with nitrosyl ligand which can release NO• under one and two-photon absorption. Compared with one-photon excitation, two-photon excitation allows high focalization and deep penetration of the beam, while exciting in the therapeutic window. This thesis is dedicated to the synthesis of [RuNO] complexes and their biological applications. The first chapter develops the state of the art in the field of ruthenium nitrosyl complexes and presents the biological issues. The second chapter presents the NO• photorelease properties of complexes with 4'-(2-fluorenyl)-2,2':6',2''-terpyridine ligand under one and two-photon excitation by spectroscopic studies. Photoproducts are characterized by X-ray diffraction. In a third chapter, cis (Cl,Cl)- and trans (Cl,Cl)-[RuII(2-fluorene-terpyridine)Cl2NO]PF6 are studied in water. The photorelease capacities of trans (NO,OH)-[RuII(fluorene-terpyridine)(Cl)(OH)(NO)]PF6 are studied in biological conditions. The fourth chapter presents the synthesis of new complexes with 4'-(2-fluorenyl)-2,2':6',2''-terpyridine ligand derivatives and their photorelease properties. The fifth chapter describes the phototoxic studies of these complexes on cancer cells (HCT 116 and FaDu). Finally, in the sixth chapter, the outstanding properties of these systems in the falling of antibiotic resistance in Staphylococcus epidermidis are exposed

Orientador: Mario Jose Abdalla Saad
Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas
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Resumo: As óxido nítrico sintases (NOS) são divididas em dois grandes grupos de enzimas, NOS induzível (iNOS) e NOS constitutivas (cNOS). Embora o óxido nítrico (NO) seja um importante mediador de defesa do organismo, a produção excessiva de NO está envolvida na patogênese de muitas doenças inflamatórias e metabólicas. Alguns estudos demonstram que o óxido nítrico exógeno e o NO produzido pela iNOS pode induzir resistência à insulina em músculo e desempenha um papel importante na hiperglicemia de jejum. Este estudo teve como objetivo sintetizar e investigar o efeito de um potente e seletivo inibidor de atividade da iNOS, o Iodato de S-Metilisotiouréia (I-SMT) 5 mg/kg por dia, na hiperglicemia de jejum e na resistência à insulina em um modelo de obesidade induzida por dieta hiperlipídica. Foram observados os parâmetros metabólicos e de sinalização celular da Proteína quinase B/Akt (Akt) e os resultados fornecem evidências de que o grupo tratado com I-SMT foi protegido contra o desenvolvimento de resistência à insulina, e intolerância à glicose induzida por dieta hiperlipídica. Portanto, propomos que potentes inibidores farmacológicos, com seletividade significativa pela iNOS podem representar uma nova abordagem terapêutica para o tratamento da resistência à insulina e suas complicações como o diabetes tipo 2.
Abstract: Nitric oxide synthase (NOS) has been divided into two major sub-enzymes, inducible NOS (iNOS) and constitutive NOS (cNOS). Although nitric oxide (NO) is an important defense mediator, the excessive production of NO has been involved in the pathology of many inflammatory and metabolic diseases. Some studies demonstrate that exogenous nitric oxide (NO) and the NO produced by iNOS can induce insulin resistance in muscle and plays an important role in fasting hyperglycemia. This study investigates the effect of a potent and selective iNOS activity inhibitor, the S-Methylisothiourea Iodide (SMT-I) 5 mg/kg per day, in fasting hyperglycemia and insulin resistance in diet-induced obesity model. We observed the metabolic parameters and Akt signalization and these findings provide evidence that the SMT-I treated group are protected against the development of insulin resistance, glucose intolerance and diet-induced obesity. Therefore, we propose that highly selective inhibitors of iNOS activity may represent a novel therapeutic approach for the therapy of insulin resistance and its complications as type 2 diabetes.
Mestrado
Biologia Estrutural, Celular, Molecular e do Desenvolvimento
Mestre em Fisiopatologia Médica

Systemic acquired resistance (SAR) is a form of inducible defense response triggered upon localized infection that confers broad-spectrum disease resistance against secondary infections. Several factors are known to regulate SAR and these include phenolic phytohormone salicylic acid (SA), phosphorylated sugar glycerol-3-phosphate (G3P), and dicarboxylic acid azelaic acid (AzA). This study evaluated a role for free radicals nitric oxide (NO) and reactive oxygen species (ROS) in SAR. Normal accumulation of both NO and ROS was required for normal SAR and mutations preventing NO/ROS accumulation and/or biosynthesis compromised SAR. A role for NO and ROS was further established using pharmacological approaches. Notably, both NO and ROS conferred SAR in a concentration dependent manner. This was further established using genetic mutants that accumulated high levels of NO. NO/ROS acted upstream of G3P and in parallel to SA. Collectively, these results suggest that NO and ROS are essential components of the SAR pathway.

Le syndrome métabolique (SMet) est caractérisé par la présence chez le même individu de plusieurs anomalies parmi les suivantes: une adiposité abdominale, une insulino-résistance (IR), une intolérance au glucose, une hypertension artérielle et une dyslipidémie. Des études ont révélé que la modulation de la voie de signalisation NO/GMPc dans le SMet peut exercer des effets métaboliques et cardiovasculaires protecteurs. Nous avons exploré, dans ce contexte, l'effet du mirabegron et du BAY 41-2272, deux molécules connues pour leur capacité à activer la voie NO-GMPc. Nous avons d'abord développé un modèle animal expérimental avec deux facteurs principaux du SMet, la dyslipidémie et l’IR. Nos résultats ont montré qu'après 12 semaines d'alimentation riche en fructose et en graisses (HFFD), le lapin Watanabe (WHHL), un modèle animal de dyslipidémie spontanée, présentait une intolérance au glucose, une IR (test HOMA-IR), une aggravation de la dyslipidémie et une diminution de la contractilité cardiaque (approche ex-vivo). Après 12 semaines de traitement, le mirabegron et le BAY 41-2272 ont prévenu le gain de poids et l’augmentation du taux de TG et ont amélioré la sensibilité à l'insuline, la fonction endothéliale des artères carotides et la fonction cardiaque (mirabegron). Ce travail a permis de mettre en place un modèle expérimental combinant la dyslipidémie et l’IR chez le lapin WHHL. De plus, les résultats ont montré que l'activation à long terme de la voie de signalisation NO-GMPc représente une approche pharmacologique prometteuse dans la gestion des complications métaboliques et cardiovasculaires associées au SMet
Metabolic syndrome (MetS) is characterized by abdominal adiposity, insulin resistance (IR), glucose intolerance, arterial hypertension and dyslipidemia. Experimental studies have revealed that modulation of the nitric oxide-cyclic guanosine monophosphate (NO-cGMP) signaling pathway in MetS can exert protective metabolic and cardiovascular effects. In this regard, we explored the effect of mirabegron and BAY 41-2272, two molecules known for their ability to activate the NO-cGMP pathway. We first developed an experimental animal model with two main components of the MetS, dyslipidemia and IR. Our results showed that after 12 weeks of high-fructose high-fat diet (HFFD) feeding, the Watanabe heritable hyperlipidemic (WHHL) rabbit, an animal model of spontaneous dyslipidemia, exhibited glucose intolerance, IR (HOMA-IR test), an aggravation in dyslipidemia and a decrease in cardiac contractility (ex-vivo approach). Twelve weeks of mirabegron and BAY 41-2272 treatment prevented weight gain and the increase in TG levels and improved insulin sensitivity, carotid endothelial function, and cardiac function (mirabegron). We were able to develop an experimental model combining dyslipidemia and IR in the WHHL rabbit. Furthermore, our results showed that long-term activation of the NO-cGMP signaling pathway represents a promising pharmacological approach in the management of the MetS and its metabolic and cardiovascular consequences

The aim of our study was to evaluate the effects of a resistance exercise session on the vascular action of insulin in superior mesenteric artery of rats. Wistar rats (250-300g) were divided into 3 groups: control (CT, n = 20), electrically stimulated (ES, n = 5) and resistance exercise (RE, n = 20). The exercise was conducted in the apparatus of the squat, where the animals were subjected to 15 sets of 10 repetitions with 3 minutes rest between sets. The intensity was set at 70% of maximum load established by repetition maximum test performed 48h before the exercise session. The animals were kept suspended in EE squat machine and received the same intensity of electrical stimuli applied to the exercised animals. Immediately after the single resistance exercise session, the animals were anesthetized and killed by exsanguination, the superior mesenteric artery was removed and sectioned into rings (1-2 mm) which were mounted on tanks for isolated organ. The endothelium-dependent relaxation was obtained from concentration-response curves for insulin on rings precontracted with phenylephrine. After that, concentration response curves were obtained for groups CT response and ER, in the absence and/or presence of the following inhibitors: L-NAME (NOS inhibitor), TEA (inhibitor nonselective channel for K+), LY294002 (PI3K inhibitor), BQ123 ( ETA receptor antagonist), and Ouabain (inhibitor of Na+/K+-ATPase), concentration-response curves to KCl in the absence and/or presence of ouabain. According to the data obtained, we found that there was no significant difference in the relaxation induced by insulin between groups EE and CT, however, the animals in the ER showed a significant increase in relaxation when compared to CT group (p<0.001). After use of L-NAME reducing the relaxation was observed in both groups (p<0.001). When evaluating the involvement of K + channels, using TEA, relaxation was inhibited only in the RE group (p <0.001). In the presence of L-NAME+TEA relaxation in the CT group was reduced and there was a group ER contraction (p<0.001). The presence of these rings LY294002 interestingly responded in similar ways in the presence of L-NAME+TEA, promoting and inhibiting the group CT contraction curve ER group (p<0.001). The BQ123 were able to amplify the relaxation in both groups (p<0.001). Using both inhibitors (LY294002+BQ123) was observed an inhibition of contractile and relaxing effects in both groups (p<0.001). To assess the functional activity of the Na+/K+-ATPase curves were made for insulin and KCl in the absence and presence of ouabain. It was observed that ouabain was a decrease of insulin-induced relaxation only in the ER (p<0.001) and reduced the relaxation promoted by KCl in both groups, this reduction being higher in the ER (p<0.001). These data demonstrate that a single session of resistance exercise promotes adjustments in insulin-induced relaxation, which is mediated by NO, the channels for K+ and the activity of Na+/K+-ATPase. In addition, there was an effect mediated by ET-1 (via ETA receptors), necessary for the control of vascular tone.
O objetivo do nosso estudo foi avaliar o efeito agudo de uma sessão de exercício resistido na ação vascular da insulina em artéria mesentérica superior de ratos. Ratos Wistar (250-300g) foram divididos em 3 grupos: controle (CT, n = 20), eletroestimulado (EE, n = 5) e exercício resistido (ER, n = 20). O exercício foi realizado no aparelho de agachamento, onde os animais foram submetidos a 15 séries de 10 repetições, com 3 minutos de intervalo entre as séries. A intensidade foi fixada em 70% da carga máxima estabelecida pelo teste de repetição máxima, realizado 48 h antes da sessão de exercício. Os animais EE foram mantidos suspensos no aparelho de agachamento e receberam a mesma intensidade de estímulo elétrico aplicadas aos animais exercitados. Imediatamente após a única sessão de exercício resistido, os animais foram anestesiados e mortos por dessangramento, a artéria mesentérica superior foi removida e seccionada em anéis (1-2 mm), os quais foram montados em cubas para órgão isolado. O relaxamento dependente de endotélio foi obtido a de curvas concentração-resposta para a insulina, em anéis pré-contraídos com fenilefrina. Após isto, foram obtidas curvas concentração-respostas para os grupos CT e ER, na ausência e/ou na presença dos seguintes inibidores: L-NAME (inibidor da NOS); TEA (inibidor não seletivo dos canais para K+), LY294002 (inibidor da PI3K); BQ123 (antagonista do receptor ETA); e Ouabaína (inibidor da Na+/K+-ATPase); curvas concentração-resposta para KCl na ausência e/ou presença de ouabaína. De acordo com os dados obtidos, constatamos que não houve diferença significativa no relaxamento induzido por insulina entre os grupos CT e EE, entretanto, os animais do grupo ER apresentaram um aumento significativo do relaxamento quando comparado ao grupo CT (p<0,001). Após a utilização do L-NAME, foi observada redução do relaxamento em ambos os grupos (p<0,001). Quando avaliamos a participação dos canais para K+, utilizando TEA foi observado inibição do relaxamento apenas no ER (p<0,001). Na presença de L-NAME+TEA o relaxamento no grupo CT foi reduzido e no grupo ER houve uma contração (p<0,001). A presença do LY294002 interessantemente estes anéis responderam de formas similares quando na presença de L-NAME+TEA, promovendo uma inibição no grupo CT e uma contração da curva no grupo ER (p<0,001). O BQ123 foi capaz de amplificar o relaxamento em ambos os grupos (p<0,001). Utilizando ambos os inibidores (LY294002+BQ123) foi observada uma inibição tanto dos efeitos contráteis quanto dos efeitos relaxantes em ambos os grupos (p<0,001). Para avaliar a atividade funcional da Na+/K+-ATPase foram feitas curvas para insulina e KCl na ausência e na presença de ouabaína. Foi observado que a ouabaína promoveu uma redução do relaxamento induzido pela insulina apenas no grupo ER (p<0,001) e reduziu o relaxamento promovidos pelo KCl em ambos os grupos, sendo esta redução maior no grupo ER (p<0,001). Estes dados demonstram que uma sessão de exercício resistido promove ajustes no relaxamento induzido por insulina, que é mediado pelo NO, pelos canais para K+ e pela atividade da Na+/K+-ATPase. Além disso, houve um efeito mediado pela ET-1 (via receptores ETA), necessária para o controle do tônus vascular, durante o exercício.

Introdução: O sedentarismo e obesidade têm sido descritos como fatores de risco relevantes para o desenvolvimento de sintomas do trato urinário inferior (STUI). No presente estudo investigamos o papel da atividade física nas alterações vesicais funcionais e moleculares induzidas por dieta hiperlipídica em ratos. Material e método: Ratos machos Wistar com oito semanas de idade foram divididos aleatoriamente em quatro grupos: 1. atividade física (AF) e dieta padrão (DP); 2. AF e dieta rica em gorduras (DHL); 3. Sedentários (SED) com DP; 4. SED e DHL. Os ratos dos Grupos 1 e 2 foram submetidos a um protocolo de treinamento de natação por 10 semanas. Ao final do protocolo, realizamos estudo funcional vesical (urodinâmico) nos ratos anestesiados, e extraímos o tecido vesical para estudo molecular da via de sinalização da insulina (IRS1/IRS2/PI3K/AKT/eNOS) através do método de reação em cadeia da polimerase quantitativa em tempo real (qRT-PCR). Resultados: Após 10 semanas, os grupos 1 e 2 apresentaram respectivamente menor ganho de peso corporal quando comparados aos grupos 3 (213,89 vs 261,63 gramas, p=0,04) e 4 (209,84 vs 257,57 gramas, p=0,04), assim como menor peso da gordura epididimária quando comparados aos grupos 3 (14,06 vs 19,74 gramas, p=0,01) e 4 (15,26 vs 20,38, p=0,02). Na soma das intervenções (sedentarismo e dieta hiperlipidica), o grupo 4 apresentou maior nível de insulina sérica (6,05 vs 4,14 ng/ml, p=0,038) e índice HOMA-IR (1,95 vs 1,09, p=0,006) em comparação ao grupo 1, sugerindo padrão de resistencia à insulina. A análise urodinâmica, na comparação do grupo 4 vs grupo 1, demonstrou padrão de hiperatividade vesical naquele: maior numero de micções (13,6 vs 6,0, p=0,04), maior pressão pós miccional (8,06 vs 5,08 mmHg, p=0,04), menor capacidade (0,29 vs 0,91 ml, p=0,008) e complacência vesical (0,027 vs 0,091ml/mmHg, p=0,016). Quanto ao estudo molecular da via de sinalização da insulina no tecido vesical, o efeito da dieta hiperlipídica resultou em subexpressão de toda a via (IRS1, IRS2, PI3K, AKT e NOS3). Em contrapartida, o efeito da atividade física resultou em superexpressão da via, em especial quando comparamos ratos com ingesta de gordura (grupo 2 x 4), assim como na comparação dos grupos \"extremos\" (grupo 1 x 4). Conclusão: A exposição dos ratos a DHL resultou em subexpressão molecular da via de sinalização de insulina no tecido vesical, assim como a AF gerou superexpressão desta, em especial nos ratos expostos a DHL. A exposição dos animais a DHL em associação a hábitos sedentários resultou em obesidade, resistência à insulina e padrão sugestivo de hiperatividade vesical em comparação aos expostos a AF e DP
Introduction: Sedentary lifestyle and obesity have been described as relevant risk factors for the development of lower urinary tract symptoms. In the present study we investigate the role of physical activity in the functional and molecular bladder alterations resulting from obesity induced by hyperlipidic diet in rats. Material and methods: Wistar male rats at eight weeks of age were randomized into groups: 1. physical activity (AF) and standard diet (DP); 2. AF and high fat diet (DHL); 3. sedentary (SED) with DP; 4.SED and DHL. Group 1 and 2 rats were subjected to a 10-week swimming training protocol. Urodynamic study was performed and expression of genes in bladder tissue (IRS1 / IRS2 / PI3K / AKT / eNOS) was evaluated. Results: Results: Groups 1 and 2 presented lower body weight gain than groups 3 (213.89 vs 261.63 grams (g), p=0.04) and 4 (209.84 vs 257.57 g, p=0.04), respectively. Group 4 had higher insulin level (6.05±1.79 vs 4.14±1.14 ng/ml, p=0.038) and higher homeostasis model assessment of insulin resistance (HOMA-IR) index (1.95 vs 1.09, p=0.006) than group 1. Group 4 had greater number of micturitions (13.6 vs 6.0, p=0.04), higher post-void pressure (8.06 vs 5.08, p=0.04), lower capacity (0.29 vs 0.91 ml, p=0.008) and lower bladder compliance (0.027 vs 0.091 ml/mmHg, p=0.016) versus group 1. HFD resulted in underexpression throughout insulin signaling pathway. Physical activity resulted in overexpression of the pathway, especially in comparisons between the rats with fat intake (groups 2 and 4) and the extreme groups (groups 1 and 4). Conclusion: Exposure of rats to hyperlipidic diet in addition to a sedentary pattern resulted in obesity, insulin resistance, urodynamic pattern suggestive of bladder hyperactivity and molecular subexpression of the IRS2 / PI3K / AKT / eNOS pathway in comparison to rats exposed to physical activity and standard diet. Physical activity generated overexpression of this molecular pathway, especially in rats exposed to a hyperlipidic diet

The direct relationship between resistance exercise and vascular health is certain, but the complex set of metabolic and hemodynamic pathways and the effects of this type of exercise on cardiovascular tissues are still controversial. Although consensus that nitric oxide (NO) contributes to the control of blood flow during resistance exercise, are not yet fully understood the events signaling in the vasculature that mediate the release of this vasoactive agent. In this sense, the objective of the present study was to evaluate the acute effects of different intensities of resistance exercise on vascular adjustments in the superior mesenteric artery of healthy rats. The thesis comprises two chapters, consisting of two original articles. The first article, "Resistance exercise enhances insulin mesenteric artery acutely-induced relaxation in healthy rats", which assessed the acute effects of an exercise session held on vascular actions of insulin in rat mesenteric artery. In this first study, it was observed that a resistance exercise session increased the vasorelaxation via PI3K/eNOS. Such increase is due in part to an increased production of NO, associated with an increase in the participation of channels for K+ and Na+/K+-ATPase. The second article, "Endothelium adjustments to acute resistance exercise are intensity-dependent in healthy animals", demonstrated that a session of vigorous exercise moderately weathered and/or improves endothelium-dependent relaxation induced by insulin due to an increase in the levels of phosphorylation of eNOSser1177 and, consequently, increased endothelial production of NO healthy animals. From these findings it is possible to suggest that resistance exercise promotes vascular adjustments that work in favor of better control of vascular tone. In addition, the magnitude of these beneficial vascular adjustments is strongly related to increased resistance exercise intensity from the intensity of 50% of 1 RM.
A relação direta entre o exercício resistido e a saúde vascular é certa, mas o complexo conjunto de vias metabólicas, hemodinâmicas e os efeitos desta modalidade de exercício físico sobre os tecidos cardiovasculares ainda é bastante controverso. Embora seja consenso na literatura que o óxido nítrico (NO) contribui para o controle do fluxo sanguíneo durante o exercício resistido, ainda não estão completamente compreendidos os eventos de sinalização na vasculatura que medeiam à liberação deste agente vasoativo. Neste sentido, o objetivo do presente estudo foi avaliar os efeitos agudos de diferentes intensidades de exercício resistido sobre os ajustes vasculares em artéria mesentérica superior de ratos saudáveis. A tese é composta por dois capítulos, constituídos de dois artigos originais. O primeiro artigo, Resistance exercise acutely enhances mesenteric artery insulin-induced relaxation in healthy rats , no qual avaliou os efeitos agudos de uma sessão de exercício resistido sobre as ações vasculares da insulina em artéria mesentérica de ratos. Neste primeiro estudo foi demonstrado que uma sessão de exercício resistido aumentou o vasorelaxamento via PI3K/eNOS. Este aumento, se deve em parte a uma maior produção de NO, associado a um aumento da participação dos canais para K+ e da Na+/K+-ATPase. O segundo artigo, Endothelium adjustments to acute resistance exercise are intensity-dependent in healthy animals demonstrou que uma sessão de exercício resistido moderado e/ou vigoroso melhora o relaxamento dependente do endotélio induzido por insulina devido a um aumento nos níveis de fosforilação da eNOS ser1177 e, consequente, incremento da produção endotelial de NO em animais saudáveis. A partir destes achados é possível sugerir que o exercício resistido é capaz de promover ajustes vasculares importantes que atuam diretamente no favorecimento do melhor controle do tônus vascular. Além disso, a magnitude destes benéficos ajustes vasculares estão fortemente relacionados ao aumento da intensidade do exercício resistido a partir da intensidade de 50% de 1 RM.

Esta tese avaliou a função em artérias de resistência do músculo esquelético (ARME) e em coronárias septais (CS) de ratos com disfunção ventricular esquerda leve (LDV) e severa (SDV) pós-infarto do miocárdio (IM). As ARME e as CS foram montadas em miógrafo de arame. O relaxamento dependente do endotélio, mediado pela acetilcolina (ACh), foi reduzido nas ARME dos SDV e preservado nos LDV. A resposta à ACh foi reduzida na CS dos SDV e aumentada nos LDV. O relaxamento mediado pelo doador de óxido nítrico (NO) foi preservado em todas as artérias. Utilizando-se a sonda DAF-2 em cortes de CS, antes e após estimulação com ACh, detectou-se menor produção de NO nos SDV e maior produção de NO nos LDV. Na CS observou-se aumento das espécies reativas de oxigênio nos SDV e redução destas nos LDV. Detectou-se maior expressão protéica da sintase de NO endotelial e neuronal e das isoformas da superóxido dismutase na CS dos LDV em relação ao SHAM. Conclui-se que a função nas ARME e nas CS é modulada diferentemente na dependência do estado da função cardíaca pós-IM.
This study assessed the reactivity in skeletal muscle resistance arteries (SMRA) and in septal coronary arteries (SC) from rats with slight (sliLVD) and severe (sevLVD) left ventricular dysfunction after myocardial infarction (MI). SMRA and the SC were mounted on a wire myograph. The endothelium-dependent relaxation by acetylcholine (ACh) was reduced in SMRA from sevLVD; but it was preserved in sliLVD. The ACh-relaxation was reduced in SC from sevLVD; while it was increased in sliLVD. The nitric oxide (NO) donor relaxation was unchanged in all arteries. Using DAF-2 probe in SC, before and after ACh stimulation, it was observed NO production decreased in SC from sevLVD, while it was increased in sliLVD. Reactive oxygen species was increased in SC from sevLVD and it was reduced in sliLVD. Neuronal and endothelial NO synthase and superoxide dismutase isoforms protein expression were greater in SC from sliLVD as compared to SHAM. In conclusion, SMRA and SC function is differentially modulated on the dependency of the cardiac function after MI.

A Arginina (Arg) regula a secreção de GH e insulina, e é o único precursor biológico do NO. Previamente demonstramos que animais tratados cronicamente com Arg (35mg/dia) desenvolvem resistência à insulina (RI), e o presente estudo investigou as suas bases moleculares. A RI baseou-se na redução da atividade e/ou expressão do IRS 1/2 e Akt, e do conteúdo de GLUT4; sendo o GH crucial na gênese desses efeitos. Doses mais elevadas de Arg (70mg/dia/30 dias), a maior geração de NO e a melhora do fluxo sangüíneo reverteram este quadro. Experimentos com células musculares demonstraram que a Arg estimula o metabolismo de glicose e lipídios, via NO/c-GMP. Esses achados indicam que a Arg pode ser benéfica para o tratamento de distúrbios metabólicos, como obesidade e DM2; e que ao estimular a secreção de GH, em doses adequadas, seria eficaz na terapia de distúrbios da secreção deste hormônio. Todavia, estudos adicionais são necessários para investigar a melhor dose e os efeitos crônicos in vivo da Arg, uma vez que o GH em excesso apresenta um efeito diabetogênico importante.
Arginine (Arg) regulates the secretion of GH and insulin, and it is the main biological precursor of NO. We have previously shown that animals chronically-treated with Arg (35 mg/day) developed insulin resistance (IR), and this study investigated its molecular basis. The RI relies on the reduction of the activity and/or expression of IRS 1/2 and Akt, and of the GLUT4 content; and GH has a crucial role in the genesis of these effects. Higher doses of Arg (70 mg/dia/30 days), the increased NO generation and the improvement of the blood flow reversed the RI. Experiments with muscle cells showed that Arg stimulates glucose and lipids metabolism, via NO/c-GMP activation. These findings indicate that Arg may be beneficial for the treatment of metabolic disorders, such as obesity and T2DM, and by stimulating GH secretion, Arg can, in appropriate doses, be effective for the therapy of GH secretion disorders. However, further studies are needed to investigate the best dose and the chronic effects of Arg in vivo, since that GH in excess is potentially diabetogenic.

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Objectives: The aim of the present study was to compare the acute effects of traditional resistance training versus power training on metabolic, cardiovascular and psychophysiological responses in elderly hypertensive women. Methods: Fifteen female elderly women (aged, 67.1 ± 6.9 years; height, 151 ± 7 cm), classified with hypertension stage 1 or 2. Subjects were randomly allocated to complete the trainings, one week later subjects allocated to RT completed the PT session and vice-versa. Training sessions were comprised of 10 exercises performed at 70% of 10 RM, involving three sets of 10 repetitions with 1 min of rest between sets and exercises. The resistance exercises for both traditional RT and PT were: barbell bench press, horizontal leg press, front lat pull-down, leg extension, military press, leg curl, cable elbow extension, standing calf raise, standing elbow flexion and dorsiflexion (Physicus, Auriflama, Sao Paulo, Brazil). Before both sessions subjects were allowed to warm-up for 10 min on a treadmill (Movement-PRO 150, Sao Paulo, Brazil) at 65-75% of heart rate reserve (HRR). During traditional RT subjects were advised to complete the concentric and eccentric phase of the movement at a moderate pace (2-3 s each), while during PT they completed the concentric phase as fast as possible and the eccentric phase with a 2-3 s cadence. Cardiovascular measures (HR and BP) were controlled during sessions to avoid any abnormal response. Heart rate, blood pressure, affective response, subjective effort perception and blood samples to analyze lactate, nitrate, nitrite, and pro- and antioxidant capacity were collected before and after training sessions. Subjects received nutritional orientation regarding nutrients that could affect cardiovascular and nitrate/nitrite analysis. Results: Systolic blood pressure was not statistically significantly different (p > 0.05) in the RT session compared with the PT at the beginning and during 30 min after sessions. Diastolic blood pressure, rate pressure product, and heart rate wereas not statistically significantly different (p > 0.05) in the RT session compared to PT at the beginning and during 45 min after sessions. Nitric oxide was statistically significantly higher (p < 0.0005) for power session as with compared to RT session after 30 min of exercise. TABARS and TROLOX were significantly higher (p < 0.05) for power session as compared with RT session only immediately after exercise. There were no differences for psychophysiological variables between protocols. Conclusion: The acute cardiovascular and metabolic response, including the oxidative stress are transient and within normal values for these variables. Taken together with the positive affective responses, both PT and RT with this intensity and volume seem to be safe for elderly hypertensive women under medication.
Objetivos: O objetivo do presente estudo foi comparar os efeitos agudos do treinamento de força tradicional versus o treinamento de potência muscular sobre as respostas metabólicas, cardiovasculares e psicofisiológicas em mulheres idosas hipertensas. Métodos: Foram selecionadas 15 mulheres idosas (idade, 67,1 ± 6,9 anos, altura, 151 ± 7 cm), classificadas com hipertensão estágio 1 ou 2. Todos os sujeitos foram alocados aleatoriamente para completar os treinos, com diferença de uma semana entre as sessões. As sessões de treinamento foram compostas por 10 exercícios realizados a 70% de 10 RM, envolvendo três séries de 10 repetições com 1 min de descanso entre séries e exercícios. Os exercícios de resistência para TP e TF tradicional foram: supino reto com barra, leg-press horizontal, puxador frontal, cadeira extensora, desenvolvimento máquina, mesa flexora, extensão dos cotovelos, flexão plantar em pé, flexão do cotovelo com barra livre e dorsiflexão (Physicus, Auriflama, São Paulo, Brasil). Antes das duas sessões, os participantes foram autorizados a aquecer por 10 min em uma esteira (Movimento-PRO 150, São Paulo, Brasil) a 65-75% da reserva de frequência cardíaca (FCR). Durante a TF tradicional, os indivíduos foram aconselhados a completar a fase concêntrica e excêntrica do movimento em um ritmo moderado (2-3 s cada), enquanto durante o TP completaram a fase concêntrica o mais rápido possível e a fase excêntrica com um 2-3 s cadência. Medidas cardiovasculares (frequência cardíaca e pressão arterial) foram controladas durante as sessões para evitar qualquer resposta anormal. A frequência cardíaca, a pressão sanguínea, a resposta afetiva, a percepção subjetiva do esforço e as amostras de sangue para analisar a capacidade de lactato, nitrato, nitrito e pró e antioxidantes foram coletadas antes e depois das sessões de treino. Os indivíduos receberam orientação nutricional em relação a nutrientes que poderiam afetar a análise cardiovascular e de nitrato / nitrito. Resultados: A pressão arterial sistólica não foi diferente (p> 0,05) na sessão TF em comparação com TP no início e durante 30 minutos após as sessões. A pressão sanguínea diastólica, o duplo produto e a frequência cardíaca não foram significativamente diferentes (p> 0,05) na sessão TF em comparação com TP no início e durante 45 minutos após as sessões. O óxido nítrico apresentou uma elevação superior (p <0,0005) para sessão de potência em comparação com a sessão de TF após 30 minutos de exercício. Thiobarbituric acid reactive substances (TABARS) e a capacidade antioxidante equivalente Trolox (TEAC) foram significativamente maiores (p <0,05) para a sessão de potência em comparação com a sessão TF apenas imediatamente após o exercício. Não houve diferenças para as variáveis psicofisiológicas entre os protocolos. Conclusão: A resposta aguda cardiovascular e metabólica, incluindo o estresse oxidativo, é transitória e dentro dos valores normais para estas variáveis. Tomados em conjunto com as respostas afetivas positivas, TP e TF com essa intensidade e volume parecem ser seguros para mulheres hipertensas idosas sob medicação.

La palourde japonaise, Ruditapes philippinarum, a été introduite en France en 1972 suite à une volonté de diversification de la production aquacole des bivalves. A la fin des années 1980, des épisodes de mortalité massive ont été observés dans les parcs vénéricoles du pays des Abers (Finistère, France). La mortalité massive des palourdes a été associée à la maladie de l’anneau brun (MAB), une vibriose dont l’agent étiologique est Vibrio tapetis. Le développement de la MAB en milieu naturel a été associé à la modulation de certains facteurs environnementaux tels que la température ou la présence de ressource trophique. Dans le cadre de ce travail et dans un premier temps, des infections expérimentales de palourdes avec différentes souches de V. tapetis ont été effectuées conjointement à une acclimatation des animaux à deux températures contrastées. L’augmentation de la température de 15 à 22 °C a été associée à de plus fortes activités enzymatiques de la superoxyde dismutase (SOD), impliquée dans le système antioxydant, et de la phénoloxydase (PO), impliquée dans le système immunitaire inné. L’augmentation de la température a également conduit à une diminution de la virulence de Vibrio tapetis. Ensemble, ces résultats permettent d’expliquer la baisse de la prévalence et de l’intensité de la MAB à 22 °C. Dans un second temps, des palourdes sexuellement matures nourries avec deux algues contrastées en termes de composition lipidique ont été infectées par V. tapetis. Si la qualité des microalgues n’a pas conduit à des différences de prévalence et d’intensité de la MAB, le statut de reproduction des animaux a eu une influence sur l’intensité de la maladie. Les réponses métaboliques de R. philippinarum exposée à V. tapetis n’ont pas été influencées par la qualité de la nourriture mais par le développement de la MAB. Ces réponses consistaient en une modulation de l’activité d’enzymes antioxydantes (SOD, catalase, glutathion peroxydase, glutathion réductase,glutathion-S-transférase) et d’enzymes reliées au système immunitaire inné (PO et oxyde nitrique synthase inductible) en fonction de la présence de signes cliniques de la MAB. Finalement, l’utilisation de ces indicateurs biochimiques pourrait permettre d’établir des critères de sélection d’individus résistants à la MAB
The Manila clam, Ruditapes philippinarum, was introduced in France in 1972 following the willingness of bivalve aquaculture diversification. In the late 1980s, episodic mass mortality events were observed in ponds of the “Pays des Abers” region (Finistère, France). The massive mortality of clams was associated brown ring disease (BRD), a vibriosis which causative agent is Vibrio tapetis. BRD development in field has been associated with the modulation of environmental factors such as temperature or the presence of trophic resource. Firstly in the frame of the present work, experimental infections of clams with different strains of V. tapetis were performed together with animal acclimation at two contrasted temperatures. The increase of temperature from 15 to 22 ° C was associated with higher enzymatic activities of superoxide dismutase (SOD), involved in the antioxidant system, and the phenoloxidase (PO), involved in the innate immune system. Temperature increase also led to a decrease in virulence of V. tapetis. Together, these results might explain the decline in BRD prevalence and intensity observed at 22 ° C. Secondly, sexually mature clams fed with two microalgal diets contrasted in terms of lipid composition were infected with V. tapetis. Although microalgae quality did not lead to any difference in BRD prevalence and intensity, the reproductive status of clams influenced BRD intensity. Metabolic responses of R. philippinarum exposed to V. tapetis were not influenced by the food quality but mainly by BRD development. These responses consisted in a modulation of the activity of antioxidant enzymes (SOD, catalase, glutathione peroxidase, glutathione reductase, glutathione S-transferase) and enzymes related to innate immune system (PO and inducible nitric oxide synthase) according to the presence of BRD clinical signs. Finally, the use of these biochemical indicators could allow for new criteria for selection of BRD resistant clams

Brain ischemia events are among the first three mortalities causes in the world associated with high treatments costs and in most cases some degree of permanent functional disability, thus it is necessary with new low cost and effective treatments. In the present study the potential neuroprotective effect of magnesium sulphate was evaluated by quantification of ischemic biomarkers: fluoride resistant acid phosphatase (FRAP), inorganic phosphate (Pi), lactate dehydrogenase (LDH) and nitric oxide (NO) in rats with global ischemia. Adult rats were anaesthetized (sodium thiopental 60mg.kg-1) and magnesium sulphate (1mmol.kg-1) (9 animals) or saline solution (NaCl 0,9%) (11 animals) was infused via jugular vein. Cortical sample was extracted 30 minutes after infusion (normoxic condition), thereafter it was induced global ischemia via respiratory arrest caused by jugular administrated muscular relaxant (Flaxedil, 500μL), and 30 minutes after this state a cortical sample of contralateral hemisphere was taken (hypoxia condition). Samples were preserved with proteases inhibitors (PMFS y NaF) and homogenized. Biomarkers were quantified within 24h from the experiment by the following spectroscopic methods: Gomori (FRAP), Ammonium molybdate (Pi), Pyruvate to lactate reduction (LDH) and Griess (NO). FRAP activity was quantified as an inflammatory biomarker for the first time in the cerebral cortex. Global ischemia increased all biomarkers concentrations of the saline group. During normoxia condition magnesium sulphate reduced Pi (P=0,0002) and LDH activity (P=0,001), while during hypoxia it reduces Pi (P= 0,0002) and LDH activity (P=0,03) compared to saline values. These results strongly suggest the cortical neuroprotectiveeffects of magnesium sulphate in global ischemia induced by respiratory arrest, by reduction of cellular acidosis and energetic deficit.

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Systemic hypertension (SH) is related to various cardiovascular events and emerged as a leading cause of death today. In the pathophysiology of hypertension occurring disorders in the autonomic nervous system that result in damage to neural mechanisms of central control of pressure, especially the baroreflex. Some of this damage is mediated by increased oxidative stress. Therapeutic approaches that reduce oxidative stress may prove effective in combating this disease. Sildenafil is a drug that acts by inhibiting the action of phosphodiesterase 5 (PDE5) and has been observed that this promotes a decrease in oxidative stress. The inhibition of PDE5 was efficient in improving vascular function and reduce blood pressure in experimental models. However, its effects on baroreflex control of blood pressure in the presence of renovascular hypertension have not yet been studied. The current study aimed to evaluate whether treatment with sildenafil is able to influence the baroreflex control of blood pressure. Wistar rats were subjected to sham surgery or inducer of renovascular hypertension (2K1C). After 5 weeks, the animals received vehicle (distilled water) or sildenafil citrate (45 mg / kg / day) for 7 days, totaling 4 groups: sham + vehicle, sham + sildenafil 2K1C + vehicle + 2K1C sildenafil. At the end of the treatment the animals were cannulated for hemodynamic measurements. They were evaluated: mean blood pressure, heart rate, spontaneous and induced baroreflex, cardiac autonomic tone and systemic oxidative stress. (CEUA-UFPB protocol no. 042/2015). Treatment with sildenafil had no significant effect in normotensive animals (121 ± 7 vs.118 ± 3 mmHg), but was effective in reducing blood pressure in 2K1C animals (139 ± 5 vs.175 ± 6 mmHg, p <0.01 ) the 2K1C animals showed reduced gain induced baroreflex (-1.93 ± 0.12) and spontaneous (-1.90 ± 0.22) compared to the sham group (-3.63 ± 0.31; - 3.95 ± 0.46). These parameters were normalized in hypertensive rats by treatment with sildenafil (-3.18 ± 0.23 -3.51 ± 0.29). Locks with atropine and propranolol showed alterations in the cardiac autonomic balance in the presence of hypertension, standardized, treatment, sympathetic tone (-24.5 ± 3 bpm, p <0.01) and vagal tone (110 ± 9 bpm, p <0.01). Sildenafil was also able to reduce systemic oxidative stress in 2K1C rats when compared to vehicle + 2K1C animals (1.04 ± 0.07 vs. 1.67 ± 0.08 nmol / ml). Treatment with sildenafil improved baroreflex control of blood pressure through the correction of the autonomic imbalance and reduction of oxidative stress.
A hipertensão arterial sistêmica (HAS) está relacionada a diversos eventos cardiovasculares e desponta como uma das principais causas de óbito atualmente. Na fisiopatologia da HAS ocorrem disfunções no sistema nervoso autônomo que resultam em prejuízo aos mecanismos neuronais de controle central da pressão, em especial ao barorreflexo. Parte deste dano é mediado pelo aumento do estresse oxidativo. Abordagens terapêuticas que reduzam o estresse oxidativo podem se mostrar eficazes no combate a esta doença. O sildenafil é uma droga que atua inibindo a ação da fosfodiesterase 5 (PDE5) e tem sido observado que este promove uma diminuição do estresse oxidativo. A inibição da PDE5 se mostrou eficiente na melhora da função vascular e redução da pressão arterial em modelos experimentais. Contudo, seus efeitos sobre o controle barorreflexo da pressão arterial na vigência de hipertensão renovascular ainda não foram estudados. O estudo atual teve como objetivo avaliar se o tratamento com sildenafil é capaz de influenciar o controle barorreflexo da pressão arterial. Foram utilizados ratos Wistar, submetidos à cirurgia sham ou indutora de hipertensão renovascular (2R1C). Após 5 semanas, os animais receberam veículo (água destilada) ou citrato de sildenafil (45mg/Kg/dia) durante 7 dias, perfazendo 4 grupos: sham + veículo, sham + sildenafil, 2R1C + veículo, 2R1C + sildenafil. Ao fim do tratamento os animais foram canulados para medidas hemodinâmicas. Foram avaliados: pressão arterial média, frequência cardíaca, barorreflexo espontâneo e induzido, tônus autonômico cardíaco e estresse oxidativo sistêmico. (CEUA-UFPB protocolo n. 042/2015). O tratamento com sildenafil não apresentou efeitos significativos nos animais normotensos (121 ± 7 vs.118± 3 mmHg), mas foi eficiente em reduzir a pressão arterial nos animais 2R1C (139 ± 5 vs.175 ± 6 mmHg, p < 0,01), Os animais 2R1C apresentaram redução do ganho do barorreflexo induzido (-1,93 ± 0,12) e espontâneo (-1,90 ± 0,22), quando comparados aos sham (-3,63 ± 0,31; -3,95 ± 0,46). Estes parâmetros foram normalizados nos animais hipertensos pelo tratamento com sildenafil (-3,18 ± 0,23; -3,51 ± 0,29). Os bloqueios com atropina e propranolol revelaram alterações no balanço autonômico cardíaco na vigência de hipertensão, normalizadas, pelo tratamento, o tônus simpático (-24,5 ± 3 bpm, p <0,01) e o tônus vagal (110 ± 9 bpm, p <0,01). O sildenafil também foi capaz de reduzir o estresse oxidativo sistêmico nos ratos 2R1C, quando comparado com os animais 2R1C + veículo (1,04 ± 0,07 vs. 1,67 ± 0,08 nmol/ml). O tratamento com sildenafil melhorou o controle barorreflexo da pressão arterial, por meio da correção do desbalanço autonômico e redução do estresse oxidativo.

A restrição crônica de sal na dieta tem sido recomendada como medida não medicamentosa do tratamento da hipertensão arterial sistêmica. Entre os efeitos observados desta medida terapêutica tem sido descrita uma redução dos valores da pressão arterial (PA), tanto em indivíduos normotensos e pacientes hipertensos, assim como em animais de laboratório. Outros efeitos observados são alterações do metabolismo de carboidratos e de lípides. Tanto em indivíduos normotensos e pacientes hipertensos, assim como em animais de laboratório foram observadas maiores concentrações de peptídeo C e de insulina, sem alteração da glicemia e com redução da captação periférica de glicose pelos tecidos, caracterizando um estado de resistência à insulina. No metabolismo de lípides, uma outra conseqüência da restrição crônica de sal é a maior concentração plasmática de colesterol total e triacilgliceróis. Apesar da demonstração dos efeitos do sal sobre o metabolismo de carboidratos e lípides em humanos e animais existem dados conflitantes na literatura, com resultados opostos a estes descritos. Para melhor compreensão deste fenômeno, foi desenvolvido um estudo em nosso laboratório, o qual demonstrou que ratos Wistar machos que receberam restrição crônica de sal na dieta apresentaram maiores insulinemias medidas durante um teste de tolerância à glicose e sem a constatação de resistência à insulina em adipócitos isolados avaliada pela EC50 Entretanto, o que não ficou esclarecido neste estudo é se este quadro era restrito ao tecido estudado ou se ocorria no animal como um todo, ou seja, se era um fenômeno generalizado. Tendo em vista estes resultados, um outro estudo foi desenvolvido em nosso laboratório. Analisando o animal intacto, foi verificado que a restrição crônica de sal em ratos Wistar estava associada a uma menor sensibilidade à insulina medida por meio de um clamp euglicêmico hiperinsulinêmico (CLAMP) em animais anestesiados. Também foi observado maior peso corpóreo (PC) e maior massa dos tecidos adiposos. No intuito de se compreender quais eram os mecanismos envolvidos na menor sensibilidade à insulina durante a restrição de sal na dieta, também foi feito o pareamento de peso entre os animais que receberam dietas hipo (HO), normo (NR) ou hipersódica (HR) e mesmo sem diferença entre os PC, os animais sob restrição salina mantiveram-se insulino resistentes. Em uma nova etapa foi feito o bloqueio do sistema renina angiotensina com o uso de captopril (inibidor da enzima conversora de angiotensina) ou losartan (antagonista do receptor ATI de angiotensina II) tendo sido verificado que o captopril melhorou a sensibilidade à insulina, o que não ocorreu com o uso de losartan. Outra conseqüência da restrição crônica de sal na dieta é a maior atividade do sistema nervoso simpático (SNS) e uma menor atividade da via L-arginina (LA) / óxido nítrico (NO). Assim, o objetivo deste estudo foi verificar se o SNS e a via LA/NO são mecanismos envolvidos na menor sensibilidade à insulina durante a restrição crônica de sal. Para o desenvolvimento deste estudo, ratos Wistar machos receberam dieta HO, NR ou HR desde o desmame aos 21 dias de vida até completarem 12 semanas. Ao tornarem-se adultos, os ratos foram submetidos ao implante cirúrgico de cateteres e três a cinco dias após era realizado um CLAMP em ratos acordados. No dia do experimento, após um jejum de seis horas se realizava a coleta de sangue e medidas metabólicas e hemodinâmicas. Neste momento, um grupo de ratos recebeu prazosin e propranolol para o bloqueio do SNS e outro grupo recebeu veículo. Um terceiro grupo de animais recebeu LA e outro grupo recebeu D-arginina (DA). Um quinto grupo de animais que consumiu apenas a dieta HR recebeu diltiazem (bloqueador de canal de cálcio). Quarenta e cinco minutos após a infusão das drogas o CLAMP foi iniciado. Foram medidos PC, glicemias, insulinemias, lípides, nitrato/nitrito (NOx), PA sistólica (PAS), diastólica (PAD) e freqüência cardíaca (FC). O PC foi maior na dieta HO do que na NR e HR. Também foi observado maior PC na dieta NR do que HR. Em situação basal observaram-se maiores valores de glicemia e insulinemia durante a restrição salina em comparação aos ratos em dieta NR e HR. A PAS e PAD foram maiores na dieta HR em comparação aos ratos em dieta NR e HO, enquanto a FC foi maior nos ratos em dieta HO em comparação àqueles sob dietas NR e HR. Também foram observadas maiores concentrações plasmáticas de colesterol total (COL) e triacilgliceróis (TAG) durante a restrição salina. Os animais controle apresentaram menor sensibilidade à insulina em comparação àqueles em dieta NR ou HR. O bloqueio do SNS corrigiu o efeito do sal sobre a captação de glicose, não sendo mais observada diferença entre as dietas. O bloqueio do SNS não influenciou a glicemia mas reduziu a maior insulinemia nos ratos em dieta HO, não sendo mais observada diferença entre as dietas quanto às insulinemias ao início do CLAMP. A infusão de LA melhorou a menor captação periférica de glicose nos ratos em dieta HO, sem influência sobre os animais tratados com dieta NR ou HR. Diltiazen não modificou a sensibilidade à insulina apesar de ter reduzido a PA em intensidade semelhante ao bloqueio do SNS. O bloqueio do SNS reduziu os valores de PAS e PAD nas três dietas, com queda mais intensa dos seus valores nos animais sob sobrecarga salina, logo ao início do CLAMP. Diltiazen reduziu a PAS e PAD. DA não influenciou (p>0,05) os valores de PAS e PAD. Já a infusão de LA promoveu redução dos valores de PAS e PAD somente em ratos em sobrecarga salina e reduziu os maiores valores de FC observados nos ratos durante a restrição de sal na dieta. O bloqueio do SNS e a infusão de LA reduziram os maiores valores de TAG ao término do CLAMP, o que não foi observado nos grupos veículo e DA. Não foi observada modificação da concentração plasmática de COL entre os grupos independentemente dos fármacos ministrados. Durante a restrição salina foi observado menor concentração plasmática de NOx em comparação à sobrecarga salina. A infusão de LA promoveu um incremento na concentração plasmática de NOx ao término do CLAMP nas três dietas, o que não foi observado nos ratos tratados com DA. Ao término do CLAMP não foram observadas diferenças na concentração de NOx plasmático entre as dietas nos animais que receberam LA. A concentração plasmática de NOx nas três dietas foi menor nos ratos que receberam DA. Assim, o bloqueio do SNS e a ativação da via LA/NO melhoraram os efeitos metabólicos decorrentes da restrição crônica de sal.
Chronic dietary salt restriction is recommended as non-pharmacological measure of hypertension treatment. One of the observed effects of this therapeutic measure is the blood pressure (BP) decrease in normotensive subjects and in hypertensive patients, and also in laboratory animals. Another side effects observed are disorders of carbohydrate and lipid metabolism. Salt restriction induced higher C-peptide and insulin levels, without changes on plasma glucose, and lower glucose uptake by the tissues in normotensive subjects and in hypertensive patients, and also in laboratory animals, suggesting an insulin resistant state. Other consequences of salt restriction are higher plasma levels of cholesterol and triacylglycerols. Some studies have disclosed opposite results. With the objective to better understand these phenomena, a study was developed in our laboratory, that showed that male Wistar rats on chronic salt restriction presented higher insulin plasma levels measured during a glucose tolerance test, without insulin resistance in isolated adipocytes measured by the EC50 of the insulin - glucose uptake curve. It was not clear in this study if this phenomenon was restricted to the evaluated tissue or if it was a phenomenon in the whole animal. With these results, another study was developed in our laboratory. Analyzing the whole animal, chronic salt restriction in male Wistar rats was associated with lower insulin sensitivity measured by a euglycemic hyperinsulinemic clamp (CLAMP) in anesthetized rats. Higher body weight (BW) and adipose tissue mass was also observed. With the objective to understand the involved mechanisms in the lower insulin sensitivity due to dietary salt restriction, weight was paired among animals on low (LSD), normal (NSD) or high (HSD) salt diet, and even without BW difference, salt restricted animals were still insulin resistants. In another step, renin angiotensin system blockade with captopril (angiotensin enzyme conversion inhibitor) or losartan (angiotensin II type I receptor antagonist) was performed. It was observed that captopril, but not losartan, improved insulin sensitivity. Another consequence of salt restriction is a higher sympathetic nervous system (SNS) and a lower L-arginine (LA) / nitric oxide (NO) pathway activity. The objective of this study was to verify if SNS and LA / NO are mechanisms involved in the lower insulin sensitivity due to chronic salt restriction. Male Wistar rats received LSD, NSD, or HSD since weaning until adulthood. In the 12th week of age, catheters were inserted and three to five days latter, a CLAMP was performed in awaked rats. On the day of the experiment, after six hours fasting, blood samples were collected and metabolic and hemodynamic measures were done. At this moment, a group of rats received prazosin and propranolol for SNS blockade and another group received vehicle. A third group of animals received LA and a fourth group received D-arginine (DA). Another group of rats only on HSD received diltiazen (calcium channel blocker). Fourty five minutes after drug infusion the CLAMP was started. BW, plasma glucose, insulin, lipids, and nitrate/nitrite (NOx), systolic BP (SBP), diastolic (DBP), and heart rate (HR) were measured. BW was higher on LSD than on NSD and HSD. BW was also higher on NSD than HSD. Basal plasma glucose and insulin were higher during salt restriction than on NSD and HSD. SBP and DBP were higher on HSD than on NSD and LSD, and HR was higher on LSD than on NSD and HSD. Cholesterol (CHOL) and triacylglycerol (TAG) plasma levels were higher on salt restriction. LSD rats presented lower insulin sensitivity compared to animals on NSD or HSD. SNS blockade corrected effect of salt on glucose uptake. SNS blockade had no influence on glucose levels but reduced the higher plasma insulin in LSD rats, without differences in insulin levels between diets at the start of the CLAMP. LA improved the lower glucose uptake observed in LSD rats, with no influence on the rats on NSD or HSD. Diltiazen had no effect on insulin sensitivity. SNS blockade reduced SBP and DBP in rats on the three diets, with an intense BP fall on HSD rats at the start of the CLAMP. Diltiazen reduced SBP and DBP. DA had no influence in SBP and DBP. On the other hand, LA decreased SBP and DBP only in salt overloaded rats and reduced the higher HR observed on salt restricted rats. SNS blockade and LA infusion reduced the higher TAG concentration at the end of the CLAMP, which was not observed in vehicle and DA groups. COL level was not influenced by drug infusion. During salt restriction, lower plasma NOx was observed compared to salt overload. LA infusion promoted plasma NOx increment at the end of the CLAMP. At the end of the CLAMP, no difference was observed in plasma NOx among the rats on the three salt diets and infused with LA. Plasma NOx was lower in rats in the DA group. In conclusion, SNS blockade and LA/NO pathway activation improved the metabolic effects due to chronic dietary salt restriction.

Hemos inducido en ratas wistar dos modelos de hipertensión arterial (HTA): hipertensión vasculorrenal 2 riñones-1 clip (2R-1C) e hipertensión por déficits crónico de óxido nítrico (NO). La angiotensina II (AII) participa en el desarrollo y mantenimiento de la hipertensión vasculorrenal y por déficit crónico de NO. La administración crónica de losartán, aún previniendo el desarrollo de la hipertensión por déficit crónico de NO, no confiere protección completa frente a las alteraciones hemodinámicas que dicha hipertensión conlleva. Los bloqueantes de los receptores de la AII, saralasina y losartán, poseen efectos hemodinámicos similares, aunque de distinta potencia, sobre los grupos de HTA. En condiciones normales, los efectos hemodinámicos del péptido natriurético auricular (PNA) no son modulados por los receptores de AII. La HTA potencia los efectos hemodinámicos del PNA por un efecto en el que intervienen los receptores de AII.
We have induced in Wistar rats two models of experimental hypertension: two kidney-one clip hypertension (2K-1C) and hypertension induced by chronic inhibition of nitric oxide production (L-NAME-induced hypertension). In both, angiotensin II (AII) participates in the development and maintenance of the hypertension. Chronic administration of losartan prevents the development of the L-NAME-induced hypertension, although it does not confer complete protection to the hemodynamic alterations that this hypertension causes. The AII receptor antagonists, saralasin and losartan, have similar hemodynamic effects on the hypertensive groups, although in different degrees. Moreover, the hemodynamic effects of the atrial natriuretic peptide (ANP) under normal conditions are not modulated by the AII receptor. However, hypertension increases the hemodynamic effects of the ANP by an effect partially due to the AII receptors

Les si alon-bn sont des materiaux susceptibles de remplacer des refractaires carbones utilises en siderurgie en particulier pour l'elaboration d'acier a faible teneur en c. Deux familles de composites, obtenues par des voies d'elaboration differentes, ont ete etudiees: si#3n#4-bn par compression a chaud et si alon-bn par frittage naturel. Dans le premier cas, un precurseur organique du bn, qui semble activer le frittage, a aussi ete utilise. Pour tous ces materiaux, nous avons etudie les procedes de fabrication, les microstructures, les proprietes mecaniques, les resistances aux chocs thermiques, a l'oxydation, a la corrosion par le fer liquide. La presence de bn, si elle est nefaste aux proprietes mecaniques, est favorable dans le cas des utilisations a haute temperature. De tous les ajouts de frittage testes: y#2o#3, nd#2o#3, la#2o#3, ceo#2, les oxydes de neodyme et de lanthane semblent conduire a une resistance a hautes temperatures legerement meilleure

Chloroplasts generate important cellular signals and synthesize diverse products. The chloroplast-localized protein, Nitric Oxide Associated-1 (NOA1), is implicated in nitric oxide (NO) accumulation and linked to the methylerythritol phosphate (MEP) pathway, but its role is undefined. I report that NOA1 is not essential for NO accumulation because the noa1 mutant accumulates NO when provided sucrose-supplemented media. Therefore, chloroplast function and fixed carbon, but not NOA1 are likely critical for plant NO accumulation. noa1 is also resistant to fosmidomycin, an inhibitor of the MEP pathway. This phenotype led to uncovering a potential link between the MEP and tetrapyrrole pathways. I report that fosmidomycin toxicity is light dependent and reduced by phytol supplementation. Downregulation of the tetrapyrrole pathway enhances fosmidomycin resistance, suggesting that reduced tetrapyrrole biosynthesis alleviates fosmidomycin toxicity. These findings reveal new insight into how impairment of the MEP pathway affects plants and the importance of metabolic balance for chloroplast function.

The identification of Nitric oxide (NO) as an endothelium-derived relaxing factor stimulated research into the physiology of this most important biological messenger, which maintains a healthy vascular endothelium and an anti-thrombotic intravascular environment. Healthy endothelial cells constantly produce NO to create ‘basal’ vasorelaxation via the classical L-arginine/sGC/cGMP activation cascade. Under physiological conditions this NO pathway is the fundamental to maintenance of normal cardiovascular health, and conversely it is the substrate for development of many cardiovascular disease states, when the balance in this system becomes impaired. Endothelial dysfunction, with the closely associated phenomenon of “NO resistance”, can affect any NO-sensitive tissues including blood vessels and platelets, and is now believed to trigger atherogenesis and thrombogenesis. Treatment of cardiovascular diseases associated with this phenomenon utilizing NO donors often has proved to be ineffective. Furthermore, treatment with organic nitrates is subject to development of nitrate tolerance, limiting efficacy of this class of agents. Several agents can ameliorate NO resistance over days or weeks, but there remains a problem in circumventing NO resistance in cardiac emergencies. In this thesis we demonstrate for the first time in humans partial circumvention of NO resistance with nitroxyl, a structural analogue of NO. Additionally, another NO sibling nitrite (NO₂⁻) has been attracting substantial interest in the last decade. Evidence has been accumulating that effects of nitrite are increased during hypoxia: - nitrite becomes a potent vasodilator and anti-aggregant when compared to normoxic environment. This is especially important in the situation of chronic tissue hypoxia or in acute vascular emergencies. Key findings from the experiments in this thesis are: 1. Nitrite is a potent vasodilator compared to GTN: in general nitrite vasodilator effects are significantly potentiated in hypoxia in human saphenous veins. However, in human internal mammary arteries, nitrite-induced vasodilation is not potentiated under hypoxia. Prolonged exposure of human saphenous vein to nitrite does not cause tolerance or cross-tolerance to GTN. Nitrite effects in saphenous veins are substantially inhibited by ODQ, suggesting that they are largely mediated by soluble guanylate cyclase. Haemoglobin, myoglobin and red blood cells significantly increase hypoxic potentiation of nitrite vasodilator effects in human saphenous veins. Hypoxic potentiation of nitrite is diminished when saphenous vein intrinsic myoglobin is blocked by ferricyanide. 2. In platelets, the anti-aggregatory effects of nitrite are markedly and selectively potentiated under hypoxia. However, nitrite is subject to “NO resistance”. Antiaggregatory actions of nitrite are more potent in venous relative to arterial blood and correlate with (greater) deoxyhaemoglobin levels. Deoxyhaemoglobin is the primary nitrite reductase in blood. We have also presented evidence that continuous generation of NO from endogenous nitrite is important in homeostasis of platelet aggregability. 3. Nitroxyl is a more potent anti-aggregant than SNP. Anti-aggregatory effects of nitroxyl are partially sGC mediated. Nitroxyl partially circumvents the phenomenon of “NO resistance” in platelets. Nitroxyl is also a potent dilator of human saphenous veins. Its effects are not NO-mediated but partially sGCmediated.
Thesis (Ph.D.) -- University of Adelaide, School of Medicine, 2015

碩士
國立清華大學
化學工程學系
102
Chemotherapy research highly prioritized overcoming the multi-drug resistance (MDR) effect in cancer cells. It might occur through different mechanisms, but it correlated with the over expression of integral membrane transporters, such as P-glycoprotein (P-gp), and exhibited a lower nitric oxide production, with resulting decrease of drug accumulation and cellular death. Irinotecan hydrochloride (CPT-11) was a general anti-microtubule drug used to treat cancer. It was also the drug efflux mediated by P-glycoprotein (P-gp) transporters. We supposed that supplying enough amount of nitride oxide that reduced the overexpression of P-gp. Thus, the resistance to CPT-11 could be reversed when MCF-7/ADR cells were incubated with nitric oxide. To solve above problem, we developed pH-responsive nitric oxide releasing poly (D,L-lactic-co-glycolic acid) hollow microparticles (PLGA HMs). Organic shell used PLGA and their aqueous core carried CPT-11, polyvinyl alcohol (PVA) and DETA-NONOate, a gas-generating agent when presented in acidic environments. SEM morphology and drug release profile provided visible evidences of the acid-responsive release of CPT-11 from PLGA HMs. MTT assay showed that our PLGA HMs could release nitric oxide in acid experiment and released nitric oxide to enhance the toxicity of CPT-11 to MCF-7/ADR cells, bringing about more MCF-7/ADR cells death. Furthermore, we created an animal model in order to proving that our PLGA HMs had greater effects on treating MCF-7/ADR tumor tissue. The results suggested that the developed PLGA HMs that could release nitric oxide to inhibit P-glycoprotein expression and reduced drug efflux in MDR cells and was a highly promising approach in chemotherapy.

v anglickém jazyce: Thesis abstract: Background and aims: Both obesity and metabolic syndrome form severe health problems in the whole world. Nevertheless the armament of pharmacotherapy for both diseases remains unsatisfactory. We aimed our work to main organs in risk of the mentioned diseases -liver and visceral fat using hepatocytes and visceral adipocytes as model. We detected 3 main metabolic and signalization activities- glycogenolysis, Nitric oxide (NO) production and transcription of inducible NO synthase (iNOS) in hepatocytes, lipolysis, NO production and iNOS transcription rate in adipocytes. We directed our interest to combination of peroxisome proliferation activator receptor γ (PPARγ) agonist, antagonist and β3 adrenergic agonist in the culture of epididymal rat adipocytes in the first part of our work. While in the second part we investigated the influence of β and α adrenergic mimetics, adrenergic blockers in the culture of rat high glycogen content hepatocytes. Methods: NO production was detected under the active agents treatments by detection of NO oxidative products NO2 and NO3 in media. Glycogenolysis was measured as free glucose rise released by hepatocytes into the media. NOS transcription level was extrapolated after comparative polymerase chain reaction with reverse...

Jia, Lin.
Thesis (Ph.D.)--Chinese University of Hong Kong, 2013.
Includes bibliographical references (leaves 124-146).
Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
Abstract also in Chinese.

碩士
國防醫學院
藥理學研究所
98
Endotoxin causes circulatory failure in sepsis which leads to tissue hypoperfusion and multiple organ failure. Nitric oxide (NO), carbon monoxide (CO) and hydrogen sulfide (H2S) assume particular importance for the regulation of vascular tone and the inflammatory response. NO and CO induce vasodilatation and H2S at lower concentrations (<100 M) induces vasoconstriction. Recently, adipose tissue has been recognized as an important secretory organ and releases mediators to regulate the vascular tone. Earlier studies addressed this issue in the rat aorta, a vessel that does not contribute to peripheral vascular resistance. By contrast, smooth muscle tone in small arteries and arterioles of the microcirculation (resistance vessels) is an important determinant of peripheral vascular resistance and, hence, blood-pressure regulation. We hypothesized that impairment of NO, CO and H2S release from perivascular adipose tissue (PVAT) in resistant mesenteric arteries may contribute to vascular hyporeactivity and hypotension in lipopolysaccharide (LPS)-induced septic rats. In the preliminary studies, we examined the effect of PVAT on mesenteric artery rings from adult male Wistar rats. Results indicate that the contractile response to phenylephrine (PE) is markedly reduced in intact vessels then that in vessels without perivascular fat. However, the anticontractile effect was not abolished by Nw-nitro-L-arginine methyl ester (L-NAME), DL-propargylglycine (PPG) and 1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one (ODQ). In this thesis, we further investigated the role of PVAT in mesenteric artery rings from rats sacrificed at 1, 2, 4, 6 hr after injection of LPS. Results indicate that the vascular hyporeactivity to PE (10-7-10-3 M) is observed in the mesenteric artery obtained from the LPS 1, 2, 4, and 6 groups. In normal condition, the PE-induced contraction was increased by L-NAME but was not affected by PPG or ODQ. In early endotoxemia, the PE-induced contraction was enhanced by L-NAME but reduced by PPG. In late endotoxemia, the PE-induced contraction was reduced by PPG while the PE-induced contraction was more enhanced by L-NAME and ODQ in intact vessels compared with vessels without perivascular fat. In conclusion, our results suggest that (1) NO may be more important than H2S and CO in the regulation of vascular tone in normal condition; (2) NO and H2S may equally contribute to the regulation of vascular tone in early endotoxemia; (3) H2S may be important in the regulation of vascular tone in late endotoxemia. In addition, NO and CO released from PVAT may be important in the regulation of vascular tone in late endotoxemia.

碩士
國立中興大學
微生物暨公共衛生學研究所
102
Salmonella is an intracellular pathogen in macrophages. This characteristic can lead the organism to more likely escape from immune clearance. Systematic infection, caused by multidrug-resistant (MDR) Salmonella, is usually associated with increased morbidity in humans and increased mortality in animals relative to non-MDR strains, and may result in difficulties during clinical treatment. It is still unclear whether MDR isolates are with higher survival in macrophages and induce less proinflammatory cytokines. This study was conducted to understand bacterial invasion and effects on survival, nitric oxide (NO) responses and proinflammatory cytokine secretion of Raw 264.7 macrophages infected by different antibiotic-resistant characteristics of Salmonella isolates. The main results indicated that nalidixic acid resistant, ciprofloxacin resistant, gryA/parC mutant and MDR strains were associated with higher invasiveness than nalidixic acid susceptible, ciprofloxacin susceptible, gryA/parC non-mutant and non-MDR strains, respectively. Macrophages after infected by ciprofloxacin susceptible and non-MDR strains were dying more rapidly than by the ciprofloxacin resistant and MDR strains during the early stage of infection (0.5 to 2 hrs), and very few cells infected by non-MDR strains could survive after 24 hrs. Lower levels of proinflammatory cytokines TNF-α and IL-6 were observed in ciprofloxacin resistant and in MDR strains. Our findings suggested that MDR isolates have higher long-term survival ability in macrophages. Moreover, macrophages infected with MDR strains secreting less proinflammatory cytokines of TNF-α and IL-6 could be less effective to clear extracellular bacteria and increase bacterial invasion.