Fernanda Viaro

Diabetes and Vascular Disease: Basic Concepts of Nitric Oxide Physiology, Endothelial Dysfunction, Oxidative Stress and Therapeutic Possibilities

The vascular manifestations associated with diabetes mellitus (DM) result from the dysfunction of several vascular physiology components mainly involving the endothelium, vascular smooth muscle and platelets. It is also known that hyperglycemia-induced oxidative stress plays a role in the development of this dysfunction. This review considers the basic physiology of the endothelium, especially related to the synthesis and function of nitric oxide. We also discuss the pathophysiology of vascular disease associated with DM. This includes the role of hyperglycemia in the induction of oxidative stress and the role of advanced glycation end-products. We also consider therapeutic strategies.

As bases experimentais da lesão por isquemia e reperfusão do fígado: revisão

Hepatic transplantation has become the main treatment for patients with terminal hepatic disease. Whatever the success of such surgery, the hepatic dysfunction associated with liver transplantation is an important cause of morbidity and mortality. Paradoxically, on restoring the blood supply, the liver is subjected to a further insult, aggravating the injury already caused by ischemia. This complex phenomenon is termed ischemia-reperfusion injury and involves endothelial cell dysfunction, leukocyte entrapment, platelet aggregation, oxidant stress and hepatic microcirculatory perfusion failure. This review discusses the physiopathlogicals mechanisms of liver ischemia-reperfusion injury.

Estudos hemodinâmicos e da função endotelial em porcas saudáveis após injeção em bolus endovenoso de azul de metileno

OBJECTIVE: Clinical benefit of methylene blue (MB) treating NO-induced vasoplegia has been reported in sepsis, systemic inflammatory response syndrome (SIRS) in cardiac surgery and anaphylactic shock, but its safety is sometimes questioned, mainly regarding its hemodynamic effects and the possibility of causing endothelium dysfunction. To examine the nitric oxide plasma levels and cardiovascular effects of the infusion of MB in vivo and its effects on endothelium-dependent and endothelium-independent in vitro vascular relaxation. METHODS: The study protocol included two experimental groups of female pigs: Group I (Control) - the animals (n=6) did not receive MB; Group II (MB) - the animals received 3 mg/kg of MB intravenous bolus infusion. After fifteen minutes of hemodynamic parameter recording the animals were sacrificed by exsanguination, and in vitro studies were conducted using segments of coronary, hepatic, superior mesenteric and renal arteries, to determine the effect of MB on the arterial endothelium function with regard to NO release. Nitric oxide plasma levels (NOx) were measured in each of the experimental groups. RESULTS: The results obtained in the present investigation were: 1) intravenous infusion of MB (3.0 mg/kg) caused no hemodynamic changes; 2) absolute and percent plasma NOx values did not differ between the experimental groups; and 3) in vitro study of vascular relaxation showed no significant difference between groups. These results show that MB intravenous infusion seems to be safe. This finding agrees with data from clinical experiments where MB was used to treat vasoplegic syndrome after cardiopulmonary bypass, systemic inflammatory response syndrome (SIRS) and anaphylaxis. These results were not unexpected because, as in healthy subjects, hemodynamics is only fine tuned and not fully under NO control; therefore, MB inhibiting guanylyl cyclase is not expected to do anything. CONCLUSION: Intravenous use of MB, at the investigated dose, did not cause any abnormal hemodynamic responses or impairment of endothelium-dependent relaxation.

Expression of nitric oxide synthases in the pathophysiology of cardiovascular diseases.

Nitric oxide (NO) is not only a potent vasodilator, it also inhibits platelet adherence and aggregation, reduces adherence of leukocytes to the endothelium, and suppresses proliferation of vascular smooth muscle cells. The knowledge of nitric oxide synthases (NOSs) is of extreme scientific importance, not only for understanding new pathophysiological mechanisms but, also as a target for therapeutic intervention 1. Therefore, the intention of this overview is to review the role of nitric oxide synthases in the physiological and pathological mechanisms of the cardiovascular system.

Plasma nitrate/nitrite (NOx) is not a useful biomarker to predict inherent cardiopulmonary bypass inflammatory response.

Abstract  Background and Aim: There were strong evidences that nitric oxide has capital importance in the progressive vasodilatation associated with varied circulatory shock forms, including systemic inflammatory response syndrome (SIRS), in patients undergoing cardiac surgeries for cardiopulmonary bypass (CPB). If CPB procedures, per se, are the inciting stimulus for inflammation, plasma nitrate/nitrite (NOx) excretion would be expected to be higher in these patients rather than in patients operated without CPB. In consequence, we hypothesized that increased levels of NOx would be predictive for vasoplegic syndrome. Methods: Thirty patients were assigned to three groups: Group 1—coronary artery bypass graft (CABG) roller pump CPB; Group 2—CABG centrifugal vortex pump CPB; and Group 3—heart valve surgery roller pump CPB. Sampling of venous blood for chemiluminescence plasma NOx dosage was achieved at the following time points: (1) before anesthesia induction; (2) after anesthesia induction; (3) before heparin infusion; (4) after heparin infusion; (5) CPB‐30 minutes; (6) CPB‐60 minutes; (7) before protamine infusion; (8) after protamine infusion; and (9) on return to the recovery area. Results: There were no intergroup differences regarding age and anesthetic regimen, and the number of arteries grafted was not different between the CABG groups. There were no NOx statistic differences, neither among the three groups of patients or among the surgery time. In addition, there was no correlation among NOx, lactate, and hemoglobin. Conclusions: Considering the inflammatory process intrinsic to CPB, this study reinforces the idea that plasma NOx is not useful as a biomarker of inflammatory response onset, which may or may not lead to SIRS and/or vasoplegic syndrome.

Forças mecânicas e veias safenas humanas: implicação na revascularização do miocárdio

Vascular endothelial cells are exposed to a variety of in vivo mechanical forces, specifically, shear stress for the blood flow, tensile stress from the compliance of the vessel wall and the hydrostatic pressure from containment of blood within inside the vasculature. Many authors studied hemodynamic, functional and morphological human saphenous veins alterations caused by these different forces with conflictant results. This review text was motivated with the specific aim of analyze literature data and some experimental data carried out in our laboratory. The adopted review subjects were: 1) Endothelial responses and gene regulation to shear stress; 2) Effects of the hydrostatic pressure in the endothelial cell morphology, gene expression of the endothelial cellular surface and proliferation of endothelial cells; 3) Effects of the traction on the human saphenous vein endothelium.

Disfunção endotelial causada pela pressão aguda de distensão em veias safenas humanas utilizadas para revascularização do miocárdio

OBJECTIVE: To study morphofunctional alterations induced by brief pressure increases in human saphenous veins utilized in coronary artery bypass grafting. METHOD: Saphenous veins of 20 patients undergoing coronary artery bypass grafting, were distributed into four experimental groups, control, 100 mmHg, 200 mmHg and 300 mmHg, and submitted to pressure distention over 15 seconds using Krebs solution. The evaluation included CD34 immunohistochemistry and an In vitro vascular reactivity study in organ chambers. RESULTS: The main experimental findings were 1) From pressures of 200 mmHg there was a tendency to reduce the CD34 expression which became statistically significant at 300 mmHg; 2) There was no impairment of the contraction and relaxation as evidenced by in vitro vascular reactivity tests. CONCLUSION: Although vascular reactivity impairment was not demonstrated in vitro, the CD34 expression, measured by imunohistochemistry, shows there is endothelium dysfunction at pressures of 300 mmHg.

Effect of arginine vasopressin on the canine epicardial coronary artery: experiments on V1-receptor-mediated production of nitric oxide

OBJECTIVE To determine whether arginine vasopressin releases endothelium-derived nitric oxide (EDNO) from the epicardial coronary artery. METHODS We studied segments of canine left circumflex coronary arteries suspended in organ chambers to measure isometric force. The coronary artery segments were contracted with prostaglandin F2alpha (2 x 10-6M) and exposed to a unique, strong arginine vasopressin concentration (10-6M) or titrated concentrations (10-9 a 10-5 M). RESULTS The unique dose of arginine vasopressin concentration (10-6M) induced transient, but significant (p<0.05), relaxation in arterial segments with endothelium, and an increase, not significant, in tension in arteries without endothelium. Endothelium-dependent relaxation to arginine vasopressin was inhibited by Ng-monomethyl-L-arginine (L-NMMA, 10-5M) or N G-nitro-L-arginine (L-NOARG) (10-4M), 2 inhibitors of nitric oxide synthesis from L-arginine. Exogenous L-arginine (10-4M), but not D-arginine (10-4M), reversed the inhibitory effect of L-NMMA on vasopressin-mediated vasorelaxation. Endothelium dependent relaxation to vasopressin was also reversibly inhibited by the vasopressin V1-receptor blocker d(CH2)5Try(Me) arginine vasopressin (10-6M) (n=6, P<0.05). CONCLUSION Vasopressin acts through V1 endothelial receptors to stimulate nitric oxide release from L-arginine.

Immunohistochemical evaluation of three nitric oxide synthase isoforms in human saphenous vein exposed to different degrees of distension pressures.

The effect of short duration and different degrees of distension pressures was investigated by means of immunohistochemistry of the three nitric oxide synthase isoforms in the human saphenous vein conventionally harvested from 20 patients submitted to coronary artery bypass graft. The human saphenous vein distal portion was divided into four segments, each one allocated to a different group. In Group I (control group), the human saphenous vein segment was not exposed to distension pressure. In Groups II, III, and IV, the human saphenous vein segment was exposed to 100, 200, and 300 mmHg of distension pressure, respectively. The distension pressures were applied and maintained with Krebs solution for 15 s. The human saphenous vein of the control group presented endothelial nitric oxide synthase and neuronal nitric oxide synthase in both endothelial and smooth muscle cells, while the inducible nitric oxide synthase appeared predominantly in the medial layer. Neither 100 nor 200 mmHg of pressurization affected the immunostaining of any nitric oxide synthase isoform. However, the human saphenous vein segments exposed to 300 mmHg of distension pressure showed a reduction in endothelial nitric oxide synthase content in the endothelium, but not in the tunica media. This lower endothelial nitric oxide synthase immunostaining in the intimal cells was associated with endothelial denudation. Therefore, we conclude that care should be taken when handling the human saphenous vein since just a few seconds of distension pressure above the normal systemic pressure can be sufficient to disrupt the endothelium reducing the amount of endothelial nitric oxide synthase and impairing the graft quality.

Endothelium-dependent and -independent hepatic artery vasodilatation is not impaired in a canine model of liver ischemia-reperfusion injury.

We investigated whether hepatic artery endothelium may be the earliest site of injury consequent to liver ischemia and reperfusion. Twenty-four heartworm-free mongrel dogs of either sex exposed to liver ischemia/reperfusion in vivo were randomized into four experimental groups (N = 6): a) control, sham-operated dogs, b) dogs subjected to 60 min of ischemia, c) dogs subjected to 30 min of ischemia and 60 min of reperfusion, and d) animals subjected to 45 min of ischemia and 120 min of reperfusion. The nitric oxide endothelium-dependent relaxation of hepatic artery rings contracted with prostaglandin F2a and exposed to increasing concentrations of acetylcholine, calcium ionophore A23187, sodium fluoride, phospholipase-C, poly-L-arginine, isoproterenol, and sodium nitroprusside was evaluated in organ-chamber experiments. Lipid peroxidation was estimated by malondialdehyde activity in liver tissue samples and by blood lactic dehydrogenase (LDH), serum aspartate aminotransferase (AST) and serum alanine aminotransferase (ALT) activities. No changes were observed in hepatic artery relaxation for any agonist tested. The group subjected to 45 min of ischemia and 120 min of reperfusion presented marked increases of serum aminotransferases (ALT = 2989 +/- 1056 U/L and AST = 1268 +/- 371 U/L; P < 0.01), LDH = 2887 +/- 1213 IU/L; P < 0.01) and malondialdehyde in liver samples (0.360 +/- 0.020 nmol/mgPT; P < 0.05). Under the experimental conditions utilized, no abnormal changes in hepatic arterial vasoreactivity were observed: endothelium-dependent and independent hepatic artery vasodilation were not impaired in this canine model of ischemia/reperfusion injury. In contrast to other vital organs and in the ischemia/reperfusion injury environment, dysfunction of the main artery endothelium is not the first site of reperfusion injury.