Óxido nítrico e sistema cardiovascular: ativação celular, reatividade vascular e variante genética / Nitric oxide and the cardiovascular system: cell activation, vascular reactivity and genetic variant / Óxido nítrico y sistema cardiovascular: activación celular, reactividad vascular y variante genética

O óxido nítrico (NO), primariamente identificado como um fator relaxante derivado do endotélio, é um radical livre atuante na sinalização de diferentes processos biológicos. A identificação das isoformas das sintases do NO (NOS) e a subsequente caracterização dos mecanismos de ativação celulares das enzimas possibilitaram tanto a compreensão de parte das interações fisiológicas como a compreensão de parte dos mecanismos de doença, na qual o NO está envolvido. A isoforma endotelial da NOS (eNOS), expressa principalmente no endotélio vascular, desempenha importante papel na regulação da reatividade vascular e no desenvolvimento e na progressão da aterosclerose. Esta revisão tem o propósito de contextualizar o leitor sobre a estrutura da eNOS e seus mecanismos de ativação celular. Tendo em vista os avanços da biologia molecular, trataremos ainda dos conhecidos mecanismos de regulação da expressão gênica e do papel de variantes no código genético da eNOS associados a fenótipos cardiovasculares. Embora se reconheça a importância do NO como molécula ateroprotetora, nossa atenção estará voltada à revisão de literatura envolvendo NO e sua participação na modulação do fenótipo de vasodilatação muscular.

Nitric oxide (NO), primarily identified as an endothelium-derived relaxing factor, is a free radical that signals different biological processes. The identification of NO synthase (NOS) isoforms and the subsequent characterization of the mechanisms of cell activation of the enzymes permitted the partial understanding of both the physiological interactions and of the mechanisms of the diseases in which NO is involved. Mainly expressed in the vascular endothelium, the endothelial NOS isoform (eNOS) plays an important role in the regulation of vascular reactivity and in the development and progression of atherosclerosis. The purpose of this review is to contextualize the reader about the eNOS structure and its mechanisms of cell activation. In view of the advances in molecular biology, we will also address the known mechanisms of gene expression regulation and the role of variants on the genetic code of eNOS associated with cardiovascular phenotypes. Although the importance of NO as an atheroprotective molecule is recognized, our focus will be the review of the literature on NO and its participation in the modulation of the muscle vasodilatation phenotype.

El óxido nítrico (NO), primariamente identificado como un factor relajante derivado del endotelio, es un radical libre actuante en la señalización de diferentes procesos biológicos. La identificación de las isoformas de las sintasas del NO (NOS) y la subsecuente caracterización de los mecanismos de activación celulares de las enzimas posibilitaron tanto la comprensión de parte de las interacciones fisiológicas como la comprensión de parte de los mecanismos de enfermedad, en la cual el NO está envuelto. La isoforma endotelial de la NOS (eNOS), expresada principalmente en el endotelio vascular, desempeña importante papel en la regulación de la reactividad vascular y en el desarrollo y en la progresión de la aterosclerosis. Esta revisión tiene el propósito de contextualizar al lector sobre la estructura de la eNOS y sus mecanismos de activación celular. Teniendo en vista los avances de la biología molecular, trataremos aun de los conocidos mecanismos de regulación de la expresión génica y del papel de variantes en el código genético de la eNOS asociados a fenotipos cardiovasculares. Aunque se reconozca la importancia del NO como molécula ateroprotectora, nuestra atención estará volcada a la revisión de literatura envolviendo NO y su participación en la modulación del fenotipo de vasodilatación muscular.

Fatores e mecanismos envolvidos na hipertrofia ventricular esquerda e o papel anti-hipertrófico do óxido nítrico: [revisão] / Factors and mechanisms involved in left ventricular hypertrophy and the anti-hypertrophic role of nitric oxide: [review]

A hipertrofia ventricular esquerda (HVE) ocorre em reposta à sobrecarga hemodinâmica relatada em várias condições fisiológicas e patológicas. Entretanto, ainda não está completamente elucidado se o estímulo primário para a hipertrofia é o estiramento mecânico do coração, fatores neuro-humorais, ou mesmo a interação de ambos. Esses fatores são traduzidos no interior da célula como alterações bioquímicas que levam à ativação de segundos (citosólicos) e terceiros (nucleares) mensageiros que irão agir no núcleo da célula, regulando a transcrição, e finalmente determinarão a expressão gênica que induza HVE. A HVE é caracterizada por alterações estruturais decorrentes do aumento das dimensões dos cardiomiócitos, da proliferação do tecido conjuntivo intersticial e da rarefação da microcirculação coronariana. Nos últimos anos, o óxido nítrico (•NO) surgiu como um importante regulador do remodelamento cardíaco, especificamente reconhecido como um mediador anti-hipertrófico. Vários estudos têm demonstrado os alvos celulares, as vias de sinalização anti-hipertrófica e o papel funcional do •NO. Portanto, a HVE parece desenvolver-se em decorrência da perda do balanço entre as vias de sinalização pró e anti-hipertróficas. Esses novos conhecimentos sobre as vias de sinalização pró e anti-hipertróficas permitirão desenvolver novas estratégicas no tratamento das HVE patológicas.

The left ventricular hypertrophy (LVH) occurs in response to the hemodynamic overload in some physiological and pathological conditions. However, it has not been completely elucidated whether the primary stimulation for the hypertrophy is the mechanical stretching of the heart, neurohumoral factors, or even the interaction of both. These factors are translated inside the cell as biochemical alterations that lead to the activation of second (cytosolic) and third (nuclear) messengers that will act in the cell nucleus, regulating transcription, and will finally determine the genic expression that induces LVH. The LVH is characterized by structural alterations due to the increase in the cardiomyocyte dimensions, the proliferation of the interstitial connective tissue and the rarefaction of the coronary microcirculation. Recently, nitric oxide (•NO) has appeared as an important regulator of cardiac remodeling, specifically recognized as an anti-hypertrophic mediator. Some studies have demonstrated the cellular targets, the anti-hypertrophic signaling pathways and the functional role of •NO. Thus, the LVH seems to develop as a result of the loss of the balance between the pro and the anti-hypertrophic signaling pathways. This new knowledge about the pro and anti-hypertrophic signaling pathways will allow the development of new strategies in the treatment of pathological LVH.

O bloqueio da síntese do óxido nítrico promove aumento da hipertrofia e da fibrose cardíaca em ratos submetidos a treinamento aeróbio / Nitric oxide synthesis blockade increases hypertrophy and cardiac fibrosis in rats submitted to aerobic training

OBJETIVO: O presente estudo avaliou as adaptações teciduais cardíacas em ratos submetidos a treinamento aeróbio, após o bloqueio da síntese de óxido nítrico (NO). MÉTODOS: Os animais (n = 48) foram divididos em quatro grupos: sedentários (grupo CONTROLE), hipertensos após administração de Ng-nitro-L-arginina metil éster durante sete dias (grupo L-NAME), treinados por meio de natação durante oito semanas (grupo TREINADO) e treinados e tratados com L-NAME na última semana (grupo TREINADO L-NAME). Em todos os animais foi registrada a pressão arterial (PA) e realizada a avaliação morfométrica cardíaca. RESULTADOS: Os grupos L-NAME e TREINADO L-NAME apresentaram-se hipertensos em relação aos demais (p < 0,05), porém a elevação da PA no grupo TREINADO L-NAME foi significativamente menor em relação ao L-NAME (p < 0,05). Os grupos TREINADO e TREINADO L-NAME apresentaram índice de peso cardíaco maior que os grupos CONTROLE e L-NAME (p < 0,05). Também apresentaram maiores índices de área cardíaca macroscópica e de fibrose cardíaca em relação aos demais (p < 0,05) e, quando comparados, o grupo TREINADO L-NAME mostrou-se significativamente superior (p < 0,05). CONCLUSÃO: O bloqueio a curto prazo da síntese de NO, em animais sedentários, induziu hipertensão, sem no entanto causar hipertrofia cardíaca. Nos animais treinados, a inibição da síntese de NO atenuou a hipertensão e promoveu hipertrofia cardíaca com aumento expressivo da fibrose miocárdica, sugerindo importante papel do NO nas adaptações teciduais cardíacas induzidas pelo treinamento físico aeróbio.

OBJECTIVE: The objective of the present study was to evaluate cardiac tissue adaptations in rats submitted to aerobic training after nitric oxide (NO) synthesis blockade. METHODS: The animals (n=48) were divided into four groups: sedentary (CONTROL group); hypertensive after administration of NG-nitro-L-arginine methyl ester for 7 days (L-NAME Group); trained for 8 weeks through swimming exercises (TRAINED Group);trained and treated with L-NAME during the last week (L-NAME TRAINED Group). All the animals were submitted to the experiment procedures for blood pressure (BP) readings and cardiac morphometric evaluation. RESULTS: In comparison to the other groups, the L-NAME and L-NAME TRAINED groups were hypertensive (p<0.05); however, BP elevation in the L-NAME TRAINED group was significantly lower than the L-NAME group (p<0.05). The heart weight indexes for the TRAINED and L-NAME TRAINED groups were higher than the CONTROL and L-NAME groups (p<0.05). Also they had presented higher rates of macroscopic cardiac area and cardiac fibrosis in relation to the rest (p<0.05); comparisons revealed that the values for the L-NAME TRAINED group were significantly higher (p<0.05) than the others. CONCLUSION: Short term NO synthesis blockade in sedentary animals induced hypertension but did not cause cardiac hypertrophy. In the trained animals, the inhibition of NO synthesis attenuated hypertension, induced cardiac hypertrophy and significantly increased myocardial fibrosis, indicating that NO plays an important role in cardiac tissue adaptations caused by aerobic exercise.

Sepsis: emerging role of nitric oxide and selectins

Sepse – um estado de infecção bacteriana sistêmica – frequentemente leva à falência múltipla de órgãos e associa-se a altos índices de mortalidade, apesar de progressos recentes no manejo de pacientes em unidades de terapia intensiva. Muitos dos efeitos maléficos associados à sepse são atribuídos a uma resposta inflamatória patologicamente ampliada que leva a recrutamento neutrofílico e ativação das moléculas de adesão do grupo das selectinas, durante as fases iniciais do processo . O óxido nítrico e sua diversas isoformas também foram implicados nas diversas manifestações vasculares da sepse como participantes diretos da toxicidade celular. Esta revisão descreve o papel das selectinas e do óxido nítrico em situações clínicas e experimentais de sepse, bem como os respectivos efeitos de processos terapêuticos de bloqueio.

Increase in rat plasma antioxidant activity after E. coli lipopolysaccharide administration

It is well recognized that the sensitivity of animals to lipopolysaccharide (LPS) endotoxin varies tremendously. And, it has been recently observed that Sprague-Dawley rats dramatically increase the activity of hepatic endogenous antioxidative enzyme systems after LPS administration. This finding suggests that the relative resistance of rats to LPS may be related to a concomitant increase in the activities of the hepatic antioxidant systems. This study was designed to examine if the above reported hepatic change in rats given LPS could be observed at the systemic level. Male Sprague-Dawley or Wistar rats, weighing 250 - 350 g, were given increasing doses (10 - 100 mg/kg) of LPS i.p. under 1.0% isoflurane anesthesia. Antioxidant capacity (AOC), blood gas analysis, and the cardiovascular parameters of the arterial blood of animals were determined over a 4 hour period following LPS administration. In addition, we studied the effect of pretreatment with the non-specific nitric oxide synthase inhibitor, L-N(G)-Nitroarginine methyl ester hydrochloride (L-NAME), given 50 mg/kg s.c. one and 24 hours before the administration of 20 mg/kg LPS i.p. in Sprague-Dawley rats. Rats given sufficiently high doses of E. coli LPS to produce behavioral effects also showed increased plasma AOCs in the early period after the administration of LPS. Similar changes were noted in Sprague-Dawley and Wistar rat strains, but at different doses that reflect their differential sensitivities to the LPS induced inflammatory response. Also, the resistance of the Sprague-Dawley strain of rats to LPS was not altered by the prior administration of L-NAME, nor was the plasma AOC altered. In conclusion, our study suggests that the rat strains are relatively resistant to develop the toxic signs of LPS in the early period after the administration of LPS, especially in Sprague-Dawley rats. Moreover, endotoxin-induced increases in plasma AOC may contribute to the rats' resistance to LPS intoxication.

Pancreatic nitric oxide and oxygen free radicals in the early stages of streptozotocin-induced diabetes mellitus in the rat

The objective of the present study was to explore the regulatory mechanisms of free radicals during streptozotocin (STZ)-induced pancreatic damage, which may involve nitric oxide (NO) production as a modulator of cellular oxidative stress. Removal of oxygen species by incubating pancreatic tissues in the presence of polyethylene glycol-conjugated superoxide dismutase (PEG-SOD) (1 U/ml) produced a decrease in nitrite levels (42 percent) and NO synthase (NOS) activity (50 percent) in diabetic but not in control samples. When NO production was blocked by N G-monomethyl-L-arginine (L-NMMA) (600 ÁM), SOD activity increased (15.21 +/- 1.23 vs 24.40 +/- 2.01 U/mg dry weight). The increase was abolished when the NO donor, spermine nonoate, was added to the incubating medium (13.2 +/- 1.32). Lipid peroxidation was lower in diabetic tissues when PEG-SOD was added (0.40 +/- 0.02 vs 0.20 +/- 0.03 nmol/mg protein), and when L-NMMA blocked NOS activity in the incubating medium (0.28 +/- 0.05); spermine nonoate (100 ÁM) abolished the decrease in lipoperoxide level (0.70 +/- 0.02). We conclude that removal of oxygen species produces a decrease in pancreatic NO and NOS levels in STZ-treated rats. Moreover, inhibition of NOS activity produces an increase in SOD activity and a decrease in lipoperoxidation in diabetic pancreatic tissues. Oxidative stress and NO pathway are related and seem to modulate each other in acute STZ-induced diabetic pancreas in the rat

Vascular changes after cardiopulmonary bypass and ischemic cardiac arrest: roles of nitric oxide synthase and cyclooxygenase

Cardiac surgery involving ischemic arrest and extracorporeal circulation is often associated with alterations in vascular reactivity and permeability due to changes in the expression and activity of isoforms of nitric oxide synthase and cyclooxygenase. These inflammatory changes may manifest as systemic hypotension, coronary spasm or contraction, myocardial failure, and dysfunction of the lungs, gut, brain and other organs. In addition, endothelial dysfunction may increase the occurrence of late cardiac events such as graft thrombosis and myocardial infarction. These vascular changes may lead to increased mortality and morbidity and markedly lengthen the time of hospitalization and cost of cardiac surgery. Developing a better understanding of the vascular changes operating through nitric oxide synthase and cyclooxygenase may improve the care and help decrease the cost of cardiovascular operations.

Neuronal and endothelial nitric oxide synthase gene knockout mice

Targeted disruption of the neuronal nitric oxide synthase (nNOS) and endothelial nitric oxide synthase (eNOS) genes has led to knockout mice that lack these isoforms. These animal models have been useful to study the roles of nitric oxide (NO) in physiologic processes. nNOS knockout mice have enlarged stomachs and defects in the inhibitory junction potential involved in gastrointestinal motility. eNOS knockout mice are hypertensive and lack endothelium-derived relaxing factor activity. When these animals are subjected to models of focal ischemia, the nNOS mutant mice develop smaller infarcts, consistent with a role for nNOS in neurotoxicity following cerebral ischemia. In contrast, eNOS mutant mice develop larger infarcts, and show a more pronounced hemodynamic effect of vascular occlusion. The knockout mice also show that nNOS and eNOS isoforms differentially modulate the release of neurotransmitters in various regions of the brain. eNOS knockout mice respond to vessel injury with greater neointimal proliferation, confirming that reduced NO levels seen in endothelial dysfunction change the vessel response to injury. Furthermore, eNOS mutant mice still show a protective effect of female gender, indicating that the mechanism of this protection cannot be limited to upregulation of eNOS expression. The eNOS mutant mice also prove that eNOS modulates the cardiac contractile response to ß-adrenergic agonists and baseline diastolic relaxation. Atrial natriuretic peptide, upregulated in the hearts of eNOS mutant mice, normalizes cGMP levels and restores normal diastolic relaxation.

Papel del oxido nitrico en la respiracion celular / Role of nitric oxide in cell respiration

El óxido nítrico (ON) sintetizado de manera constitutiva en una célula generadora, actúa sobre otra célula efectora como regulador fisiológico del tono vascular, de la inhibición planquetaria o de la interación neurona-neurona. En forma contraría, la liberación inducida de grandes cantidades de ON por períodos largos transforma al ON de un mediador fisiológico en una molécula citostática y citotóxica. La presencia de ON sintetasa en la mitocondria sugiere que la producción fisiológica de ON en pequeñas cantidades estaría relacionada con la regulación de la respiración celular a través de la inhibición de la citocromo oxidasa. La exposición celular a ON por períodos prolongados da como resultado una inhibición irreversible de la respiración celular que es independiente de la formación generalizada de superóxido o de peroxinitrito. La inhibición de la respiración celular es persistente excepto si se estudia el complejo IV aislado, si se bloquea el complejo I o si se pone glutatión. De esta manera nuestra hipótese es que la inhibición del complejo IV es un efecto fisiológico normal, dependiente de las concentraciones de ON. Si se expone las células a ON por períodos largos, se nitrosilan tioles en el complejo I y a medida que se están nitrosilando, el glutatión transnitrosila y limpia el complejo I hasta que el glutation cae a un nivel crítico. En estas condiciones se produce la inhibición irreversible de la respiración y posiblemente, este sea el camino que transforma al ON de un mediador fisiológico en una molécula con efectos patológicos. Asi mismo, creemos que la nitrosilación de tioles y la transnitrosilación por el glutatión constituyen un mecanismo crítico de la prevención del stress oxidativo.

Differential densities of nitric oxide synthesizing nerves in the sphincteric and non-sphincteric smooth muscles of human gut during fetal development

Nitric oxide (NO) is involved in the inhibitory neurotransmission in the sphincter and non-sphincteric smooth muscles. However, the relative contribution of nitric oxid synthesizing innervation to these functionally diverse parts of the gut, particularly during development, is unknown. Gastrointestinal sphincters and adjoining non-sphincteric bowel segments were obtained by nicotinamide adenine deoxinucleotide ages between 12 and 23 weeks. NO synthesizing nerves were examined by nicotinamide adenine deoxinucleotide phosphate (NADPH) diaphorase histochemistry. The densities of NADPH positive nerves in the smooth muscle were quantified using a computerized image analyzing system on randon section. The NO synthesizing nerve density in intestinal smooth muscles decreased during fetal development as a results of increased interspacing between myenteric ganglia and a disproportionately larger increase in smooth muscle area than neuronal area. Similarly, the nerve densities were lower in sphincteric regions than adjoining non-sphincteric regions at the same gestational ages. There is a relative reduction of the density of NO synthesizing nerves in intestinal smooth muscle particularly in sphincteric regions during development. These findings may have relevance to the occurence of congenital dysmotility disordere of the sphincteric regions