RuBPY decreases intracellular calcium by decreasing influx and increasing storage.

cis-[Ru(bpy)2(py)NO2](PF6) (RuBPY) is a ruthenium complex nitric oxide (NO) donor that presents a nitrite in its moiety and has been shown to induce vasodilation in various arteries, as well as arterial pressure reduction with no changes in heart rate. Because vascular tone is highly dependent on the cytosolic calcium concentration ([Ca2+ ]c), the current study aimed to investigate the effects of RuBPY on the intracellular mobilization of calcium stores of rat aortic vascular smooth muscle cells. Vascular reactivity experiments were performed in isolated aortic rings that were contracted with a high concentration of KCl or phenylephrine (Phe). Moreover, primary cultured vascular smooth muscle cells were used to measure [Ca2+ ]c by confocal microscopy. The NO donor RuBPY decreased the [Ca2+ ]c and reduced KCl and Phe-induced contractile responses. The selective inhibitor of sarco-endoplasmic Ca-ATPase (SERCA) with thapsigargin impaired the effect of RuBPY on Phe-induced contractile response. RuBPY also reduced caffeine-induced contraction, and the contraction dependent on the capacitive Ca2+ influx. Therefore, our results suggest that NO released from RuBPY decreased [Ca2+ ]c by calcium influx blockade and activation of guanylyl-cyclase-cGMP-GK pathway. These results indicate that RuBPY increases Ca2+ storage in the sarcoplasmic reticulum by SERCA activation and also by capacitive Ca2+ influx inhibition, which is dependent on the intracellular release of nitric oxide from this compound.

C-type natriuretic peptide-induced relaxation through cGMP-dependent protein kinase and SERCA activation is impaired in two kidney-one clip rat aorta.

AIMS: Hypertension underlies endothelial dysfunction, and activation of vasorelaxation signaling with low dependence on nitric oxide (NO) represents a good alternative for vascular modulation. C-type natriuretic peptide (CNP) causes relaxation by increasing cyclic guanosine 3',5'-monophosphate (cGMP) or Gi-protein activation through its natriuretic peptide receptor-B or -C, respectively. We have hypothesized that CNP could exerts its effects and could overcome endothelial dysfunction in two kidney-one clip (2K-1C) hypertensive rat aorta. Here, we investigate the intracellular signaling involved in CNP effects in hypertension. MATERIALS AND METHODS: The 2K-1C hypertension was induced in male Wistar rats (200 g). CNP-induced vascular relaxation and cGMP production were investigated in rat thoracic aortas. The natriuretic peptide receptor-B and -C localization was evaluated by immunofluorescence. Calcium mobilization was assessed in endothelial cells from rat aortas. KEY FINDINGS: CNP induced similar relaxation in normotensive and 2K-1C hypertensive rat aortas, which increased after endothelium removal. CNP-induced relaxation involved natriuretic peptide receptor-B and -C activation in 2K-1C rats. Nitric oxide synthase (NOS) and soluble guanylyl cyclase (sGC) counter-regulated CNP-particulate GC (pGC) activation in aortas. CNP reduced endothelial calcium and increased cGMP production, which was lower in 2K-1C. CNP-induced cGMP-dependent protein kinase (PKG) and sarcoplasmic/endoplasmic reticulum Ca2+-ATPase (SERCA) activation was impaired in 2K-1C rat aorta. SIGNIFICANCE: Our results indicated CNP triggered relaxation through its natriuretic peptide receptor-B and -C in 2K-1C rat aortas, and that CNP-induced relaxation overcomes endothelial dysfunction in hypertension. In addition, NOS and sGC activities counter-regulate CNP-pGC activation to induce vascular relaxation.

Central Administration of Angiotensin-(1-7) Improves Vasopressin Impairment and Hypotensive Response in Experimental Endotoxemia.

Angiotensin-(1-7) [Ang-(1-7)]/Mas receptor is a counter-regulatory axis that counteracts detrimental renin-angiotensin system (RAS) effects, especially regarding systemic inflammation, vasopressin (AVP) release, and hypothalamic-pituitary-adrenal (HPA) activation. However, it is not completely understood whether this system may control centrally or systemically the late phase of systemic inflammation. Thus, the aim of this study was to determine whether intracerebroventricular (i.c.v.) administration of Ang-(1-7) can modulate systemic inflammation through the activation of humoral pathways in late phase of endotoxemia. Endotoxemia was induced by systemic injection of lipopolysaccharide (LPS) (1.5 mg/kg, i.v.) in Wistar rats. Ang-(1-7) (0.3 nmol in 2 µL) promoted the release of AVP and attenuated interleukin-6 (IL-6) and nitric oxide (NO) levels but increased interleukin-10 (IL-10) in the serum of the endotoxemic rats. The central administration of Mas receptor antagonist A779 (3 nmol in 2 µL, i.c.v.) abolished these anti-inflammatory effects in endotoxemic rats. Furthermore, Ang-(1-7) applied centrally restored mean arterial blood pressure (MABP) without affecting heart rate (HR) and prevented vascular hyporesponsiveness to norepinephrine (NE) and AVP in animals that received LPS. Together, our results indicate that Ang-(1-7) applied centrally promotes a systemic anti-inflammatory effect through the central Mas receptor and activation of the humoral pathway mediated by AVP.

Testosterone increases bradykinin-induced relaxation in the coronary bed of hypertensive rats.

Physiological or supraphysiological levels of testosterone appear to be associated with the development of risk factors for cardiovascular diseases such as hypertension, as this hormone modulates the release of endothelial factors. However, its actions are still controversial, especially in the coronary circulation of hypertensive animals. This study was designed to assess the effects of testosterone treatment (T) on endothelium-dependent coronary vascular reactivity in orchiectomized SHR. The animals were divided into SHAM, orchiectomized (ORX), ORX+T and ORX+T+aromatase inhibitor (AI). All treatments lasted 15 days. Blood pressure (BP) was measured. Dose-response curves to bradykinin (BK) were constructed using the Langendorff technique, followed by inhibition of endothelium mediators (NO, prostanoids, EETs) and potassium channels. The intensity of eNOS, COX-1, COX-2, Akt, and gp91phox protein expression was quantified by Western blotting. BP was elevated in SHAM, ORX+T, and ORX+T+AI groups. However, we did not observe differences in the ORX group. Baseline coronary perfusion pressure (CPP) remained unaffected. Orchiectomy did not change the BK-induced relaxation compared to the SHAM group, whereas testosterone treatment increased it. This response was diminished in the absence of NO, prostanoids, and EETs in the SHAM and ORX groups, while in ORX+T group the relaxation was diminished only in the absence of NO and EETs. There was no difference in eNOS, COX-1, COX-2, and gp91phox protein expression, though Akt expression was increased in ORX and ORX+T groups. These results show that testosterone treatment can modulate endothelial function, especially in the coronary circulation under hypertension conditions, via NO and EETs pathways.

Reversion of cardiovascular remodelling in renovascular hypertensive 2K-1C rats by renin-angiotensin system inhibitors.

OBJECTIVES: Evaluate whether the RAS dual blockade would induce additional beneficial effects on cardiovascular remodelling when compared to monotherapy in renal hypertensive two kidneys-one clip (2K-1C) rats. METHODS: Hypertensive 2K-1C and normotensive (2K) rats were treated for 14 days with submaximal doses of losartan (LOS), enalapril (ENA), losartan plus enalapril (LOS + ENA) or vehicle (water). Blood pressure and some parameters of cardiovascular remodelling were evaluated. RESULTS: Systolic blood pressure (SBP) was higher in 2K-1C (209 ± 3 mm Hg, P < .05) than in 2K (113 ± 1 mm Hg) rats. There was an additional effect in 2K-1C treated with LOS + ENA (153 ± 9 mm Hg) on lowering SBP when compared to LOS (184 ± 12 mm Hg) or ENA (177 ± 9 mm Hg). None of the treatments had effect on SBP in 2K rats. In 2K-1C, cardiomyocyte hypertrophy was reduced by all treatments, although the cardiac hypertrophy indexes remained unchanged. 2K-1C aortas presented medial thickening that was partially reduced by the treatments. Intimal hyperplasia observed in 2K-1C (15.56 ± 0.89 µm vs 8.24 ± 0.80 µm) was reversed by ENA (9.52 ± 0.45 µm) or LOS + ENA (8.17 ± 0.53 µm). Collagen deposition was increased in 2K-1C hearts (1.77 ± 0.16 vs 1.28 ± 0.09) and aortas (8.1 ± 0.6 vs 5.2 ± 0.2). Treatment with LOS reduced (1.12 ± 0.14%) and ENA (0.81 ± 0.11%) or LOS + ENA (0.86 ± 0.11%) additionally diminished collagen only in 2K-1C hearts. CONCLUSIONS: Submaximal doses of ACEi and/or ARB have inhibitory actions on cardiac remodelling and vascular hypertrophy not entirely dependent on their effects on blood pressure normalization in renovascular hypertensive rats. Combined therapy produced additional reduction in blood pressure than monotherapy despite a similar inhibition on cardiovascular remodelling.

Central angiotensin-(1-7) attenuates systemic inflammation via activation of sympathetic signaling in endotoxemic rats.

Angiotensin-(1-7) [Ang-(1-7)] is an angiotensin-derived neuropeptide with potential anti-hypertensive and anti-inflammatory properties. However, a possible action of Ang-(1-7) in neuroimmune interactions to regulate inflammatory response has not been explored. Thus, the aim of this study was to determine whether the intracerebroventricular (i.c.v.) administration of Ang-(1-7) can modulate systemic inflammation via sympathetic efferent circuits. Wistar male rats received systemic administration of lipopolysaccharide (LPS) (1.5 mg/Kg). Ang-(1-7) (0.3 nmol in 2 µL) promoted the release of splenic norepinephrine and attenuated tumor necrosis factor (TNF) and nitric oxide (NO), but increased interleukin-10 (IL-10), levels in the serum, spleen, and liver in endotoxemic rats. Furthermore, 6-hydroxydopamine-induced chemical sympathectomy (100 mg/Kg, intravenous) or i.c.v. administration of Mas receptor antagonist A779 (3 nmol in 2 µL) abolished the anti-inflammatory effects of central Ang-(1-7) injection. Moreover, this treatment did not alter the plasmatic LPS-induced corticosterone and vasopressin. The administration of Ang-(1-7) reverted the low resistance in response to catecholamines of rings of thoracic aorta isolated from endotoxemic rats, treated or not, with this peptide by a mechanism dependent on the regulation of NO released from perivascular adipose tissue. Together, our results indicate that Ang-(1-7) regulates systemic inflammation and vascular hyporesponsiveness in endotoxemia via activation of a central Mas receptors/sympathetic circuits/norepinephrine axis and provide novel mechanistic insights into the anti-inflammatory Ang-(1-7) properties.

Effects of progesterone treatment on endothelium-dependent coronary relaxation in ovariectomized rats.

AIM: Although progesterone (P4) has a beneficial effect on the cardiovascular system, P4 actions on the coronary bed have not yet been fully elucidated. This study evaluated the effect of progesterone treatment on endothelium-dependent coronary vascular reactivity in Wistar rats. MAIN METHODS: Eight-week-old adult rats were divided into Sham, Ovariectomized (OVX), Ovariectomized and progesterone treated (OVX P4). The OVX P4 group received daily doses of progesterone (2 mg/kg/day). Vascular reactivity was assessed by a modified Langendorff technique. The intensity of eNOS, Akt, and gp91phox protein expression was quantified by Western blotting. Superoxide anion (O2●-) and hydrogen peroxide (H2O2) production was measured by dihydroethidium and 2',7'-dichlorofluorescein, respectively. KEY FINDINGS: Treatment with P4 was able to prevent the reduction in baseline coronary perfusion pressure induced by ovariectomy. We observed that endothelium-dependent coronary vasodilation was reduced in the OVX group and potentiated in the OVX P4 group. Following the inhibition of the nitric oxide (NO) pathway, the bradykinin-induced relaxing response was potentiated in the OVX P4 group. With regard to the combined inhibition of NO and prostanoids pathways, the OVX P4 group showed a greater relaxing response, similar to what was found upon individual inhibition of NO. After the combined inhibition of NO, prostanoids and epoxyeicosatrienoic acids' pathways, the vasodilatory response induced by BK was abolished in all groups. SIGNIFICANCE: Treatment with P4 prevented oxidative stress induced by ovariectomy. These results suggest that progesterone has a beneficial action on the coronary vascular bed.

The Role of Glycocalyx and Caveolae in Vascular Homeostasis and Diseases.

This review highlights recent findings about the role that endothelial glycocalyx and caveolae play in vascular homeostasis. We describe the structure, synthesis, and function of glycocalyx and caveolae in vascular cells under physiological and pathophysiological conditions. Special focus will be given in glycocalyx and caveolae that are associated with impaired production of nitric oxide (NO) and generation of reactive oxygen species (ROS). Such alterations could contribute to the development of cardiovascular diseases, such as atherosclerosis, and hypertension.

Exposure to acetaminophen impairs vasodilation, increases oxidative stress and changes arterial morphology of rats.

Acetaminophen (APAP) is one of the most widely consumed drugs in the world. Studies have shown renal and hepatic damage as the direct result of high oxidative stress induced by APAP. Since the cardiovascular system is sensitive to oxidative stress and literature describes increased cardiovascular dysfunction in APAP consumers, this work aimed to evaluate harmful effects of APAP on the vascular system. Rats were exposed to APAP (400 mg/kg/day in drinking water) for 14 days. Plasma and aortas were collected and stored in - 80 °C and a selection of arteries was prepared for isometric tension recordings, morphological, immunohistochemical and protein expression analysis. The APAP-treated group presented increased transaminases (ALT/AST) and malondialdehyde levels in the plasma compared to controls. Lipid peroxidation, glutathione reductase and superoxide dismutase levels were increased in the plasma and arteries of the APAP group. Nevertheless, glutathione level was reduced as compared to control group. The vasodilation response to acetylcholine and sodium nitroprusside (0.1 nM to 10 µM) was also impaired after APAP treatment; however, the vascular relaxation was restored after treatment with vitamin C (100 µM). Arteries from the APAP group presented reduced wall thickness, collagen deposition, elastic fibers and increased immunoreactivity to nitrotyrosine. eNOS and sGC protein expression remained unchanged and were at similar levels as controls. These findings showed higher oxidative stress and impaired vasodilation in rats exposed to APAP. Furthermore, arteries presented reduced cell layers, collagen, elastin deposition and significantly increased immunoreactivity to nitrotyrosine after APAP treatment.

LmrBPP9: A synthetic bradykinin-potentiating peptide from Lachesis muta rhombeata venom that inhibits the angiotensin-converting enzyme activity in vitro and reduces the blood pressure of hypertensive rats.

Bradykinin-potentiating peptides (BPPs) are an important group of toxins present in Lachesis muta rhombeata venom. They act directly at renin-angiotensin-aldosterone system, through the inhibition of angiotensin-converting enzyme (ACE). This action may contribute to the hypotensive shock observed during the envenoming by this species. Thus, the main goal of this study was the solid-phase synthesis of a BPP found in L. m. rhombeata venom and its in vitro and in vivo characterization in relation to ACE inhibition and hypotensive activity, respectively. The LmrBPP9 peptide was synthesized using an automated solid-phase peptide synthesizer and purified by reversed-phase fast protein liquid chromatography (FPLC). The in vitro IC50 of the synthetic peptide is 4.25 ±â€¯0.10 µM, showing a great capacity of ACE inhibition. The in vivo studies showed that LmrBPP9 induces blood pressure reduction, both in normotensive and hypertensive rats, being more pronounced in the last ones. These results agree with the in vitro results, showing that the synthetic peptide LmrBPP9 is a potential molecule to the development of a new antihypertensive drug.

GPER modulates tone and coronary vascular reactivity in male and female rats.

Compared with age-matched men, premenopausal women are largely protected from coronary artery disease, a difference that is lost after menopause. The effects of oestrogens are mediated by the activation of nuclear receptors (ERα and ERß) and by the G protein-coupled oestrogen receptor (GPER). This study aims to evaluate the potential role of GPER in coronary circulation in female and male rats. The baseline coronary perfusion pressure (CPP) and the concentration-response curve with a GPER agonist (G-1) were evaluated in isolated hearts before and after the blockade of GPER. GPER, superoxide dismutase (SOD-2), catalase and gp91phox protein expression were assessed by Western blotting. Superoxide production was evaluated 'in situ' via dihydroethidium fluorescence (DHE). GPER blockade significantly increased the CPP in both groups, demonstrating the modulation of coronary tone by GPER. G-1 causes relaxation of the coronary bed in a concentration-dependent manner and was significantly higher in female rats. No differences were detected in GPER, SOD-2 and catalase protein expression. However, gp91phox expression and DHE fluorescence were higher in male rats, indicating elevated superoxide production. Therefore, GPER plays an important role in modulating coronary tone and reactivity in female and male rats. The observed differences in vascular reactivity may be related to the higher superoxide production in male rats. These findings help to elucidate the role of GPER-modulating coronary circulation, providing new information to develop a potential therapeutic target for the treatment of coronary heart disease.

Hypotensive effect and vascular relaxation in different arteries induced by the nitric oxide donor RuBPY.

NO donors are compounds that release NO that can be used when the endogenous NO bioavailability is impaired. The compound cis-[Ru(bpy)2(py)(NO2)](PF6) (RuBPY) is a nitrite-ruthenium, since it has a NO2 in its molecule. The aim of the present study was to evaluate the effect of RuBPY on arterial pressure, as well as on the vascular relaxation of different vascular arteries in renal hypertensive (2K-1C) and normotensive (2K) rats. We have evaluated the arterial pressure and heart rate changes as well as the RuBPY and SNP-induced relaxation (thoracic aorta, mesenteric resistance, coronary and basilar arteries). The administration of RuBPY in awake rats evoked a smaller but long lasting hypotensive effect when compared to SNP, with no increase in heart rate. The relaxation induced by RuBPY was similar between 2K-1C and 2K rats in thoracic aorta, mesenteric resistance and coronary arteries. However, the relaxation induced by RuBPY was smaller in basilar arteries from 2K-1C than in 2K. Taken together, our results show that RuBPY presents several advantages over SNP, since it does not induce hypotensive effect in normotensive animals, the hypotensive effect is slower, with no reflex tachycardia, and it is long lasting. In addition, RuBPY induces coronary artery relaxation (useful for angina) and presented only a small effect on basilar artery (may not induce headache).

A Novel Vasoactive Proline-Rich Oligopeptide from the Skin Secretion of the Frog Brachycephalus ephippium.

Proline-rich oligopeptides (PROs) are a large family which comprises the bradykinin-potentiating peptides (BPPs). They inhibit the activity of the angiotensin I-converting enzyme (ACE) and have a typical pyroglutamyl (Pyr)/proline-rich structure at the N- and C-terminus, respectively. Furthermore, PROs decrease blood pressure in animals. In the present study, the isolation and biological characterization of a novel vasoactive BPP isolated from the skin secretion of the frog Brachycephalus ephippium is described. This new PRO, termed BPP-Brachy, has the primary structure WPPPKVSP and the amidated form termed BPP-BrachyNH2 inhibits efficiently ACE in rat serum. In silico molecular modeling and docking studies suggest that BPP-BrachyNH2 is capable of forming a hydrogen bond network as well as multiple van der Waals interactions with the rat ACE, which blocks the access of the substrate to the C-domain active site. Moreover, in rat thoracic aorta BPP-BrachyNH2 induces potent endothelium-dependent vasodilatation with similar magnitude as captopril. In DAF-FM DA-loaded aortic cross sections examined by confocal microscopy, BPP-BrachyNH2 was found to increase the release of nitric oxide (NO). Moreover, BPP-BrachyNH2 was devoid of toxicity in endothelial and smooth muscle cell cultures. In conclusion, the peptide BPP-BrachyNH2 has a novel sequence being the first BPP isolated from the skin secretion of the Brachycephalidae family. This opens for exploring amphibians as a source of new biomolecules. The BPP-BrachyNH2 is devoid of cytotoxicity and elicits endothelium-dependent vasodilatation mediated by NO. These findings open for the possibility of potential application of these peptides in the treatment of endothelial dysfunction and cardiovascular diseases.

In vitro Treatment with cis-[Ru(H-dcbpy-)2(Cl)(NO)] Improves the Endothelial Function in Aortic Rings with Endothelial Dysfunction.

PURPOSE: The ruthenium complex cis-[Ru(H-dcbpy-)2(Cl)(NO)] (DCBPY) is a nitric oxide (NO) donor and studies suggested that the ruthenium compounds can inactivate O2-. The aim of this study is to test if DCBPY can revert and/or prevent the endothelial dysfunction. METHODS: Normotensive (2K) and hypertensive (2K-1C) wistar rats were used. To vascular reactivity study, thoracic aortas were isolated, rings with intact endothelium were incubated with: DCBPY: 0.1; 1 and 10µM, DCBPY plus hydroxocobalin (NO scavenger) or tempol during 30 minutes, and concentration effect curves to acetylcholine were performed. The potency values (pD2) and maximum effect (ME) were analyzed. The O2- was generated using hypoxantine xantine oxidase and the reduction of cytochrome c, NO consumption by O2- and the effect in avoid NO consumption was measured. RESULTS: In 2K-1C DCBPY at 0.1; 1 or 10µM improved the relaxation endothelium dependent induced by acetylcholine in aortic rings compared to control 2K-1C, and also improved ME. In rings from 2K incubation with DCBPY (0.1; 1.0 and 10 µM) did not change pD2 or ME. Incubation with 0.1 µM of DCBPY plus hydroxocobalamin did not modify the potency and ME in 2K-1C compared to DCBPY (0.1 µM). DCBPY and SOD inhibits the reduction of cytochrome c and inhibited the NO consumption by O2-, showing that O2- has been removed from the solution. CONCLUSION: Our results suggest that DCBPY at a lower concentration (0.1 µM) is not an NO generator, but can inactivate superoxide and improves the endothelial function.

Ruthenium complexes as NO donors for vascular relaxation induction.

Nitric oxide (NO) donors are substances that can release NO. Vascular relaxation induction is among the several functions of NO, and the administration of NO donors is a pharmacological alternative to treat hypertension. This review will focus on the physicochemical description of ruthenium-derived NO donor complexes that release NO via reduction and light stimulation. In particular, we will discuss the complexes synthesized by our research group over the last ten years, and we will focus on the vasodilation and arterial pressure control elicited by these complexes. Soluble guanylyl cyclase (sGC) and potassium channels are the main targets of the NO species released from the inorganic compounds. We will consider the importance of the chemical structure of the ruthenium complexes and their vascular effects.

Calcium influx inhibition is involved in the hypotensive and vasorelaxant effects induced by yangambin.

The pharmacological effects on the cardiovascular system of yangambin, a lignan isolated from Ocotea duckei Vattimo (Lauraceae), were studied in rats using combined functional and biochemical approaches. In non-anaesthetized rats, yangambin (1, 5, 10, 20, 30 mg/kg, i.v.) induced hypotension (-3.5 ± 0.2; -7.1 ± 0.8; -8.9 ± 1.3; -14 ± 2.3, -25.5% ± 2.6%, respectively) accompanied by tachycardia (5.9 ± 0.5; 5.9 ± 1.6; 8.8 ± 1.4; 11.6, 18.8% ± 3.4%, respectively). In isolated rat atria, yangambin (0.1 µM-1 mM) had very slight negative inotropic (Emax = 35.6% ± 6.4%) and chronotropic effects (Emax = 10.2% ± 2.9%). In endothelium-intact rat mesenteric artery, yangambin (0.1 µM-1 mM) induced concentration-dependent relaxation (pD2 = 4.5 ± 0.06) of contractions induced by phenylephrine and this effect was not affected by removal of the endothelium. Interestingly, like nifedipine, the relaxant effect induced by yangambin was more potent on the contractile response induced by KCl 80 mM (pD2 = 4.8 ± 0.05) when compared to that induced by phenylephrine. Furthermore, yangambin inhibited CaCl2-induced contractions in a concentration-dependent manner. This lignan also induced relaxation (pD2 = 4.0 ± 0.04) of isolated arteries pre-contracted with S(-)-Bay K 8644. In fura-2/AM-loaded myocytes of rat mesenteric arteries, yangambin inhibited the Ca2+ signal evoked by KCl 60 mM. In conclusion, these results suggest that the hypotensive effect of yangambin is probably due to a peripheral vasodilatation that involves, at least, the inhibition the Ca2+ influx through voltage-gated Ca2+ channels.

Decreased endothelial nitric oxide, systemic oxidative stress, and increased sympathetic modulation contribute to hypertension in obese rats.

We investigated the involvement of nitric oxide (NO) and reactive oxygen species (ROS) on autonomic cardiovascular parameters, vascular reactivity, and endothelial cells isolated from aorta of monosodium glutamate (MSG) obese rats. Obesity was induced by administration of 4 mg/g body wt of MSG or equimolar saline [control (CTR)] to newborn rats. At the 60th day, the treatment was started with N(G)-nitro-L-arginine methyl ester (L-NAME, 20 mg/kg) or 0.9% saline. At the 90th day, after artery catheterization, mean arterial pressure (MAP) and heart rate were recorded. Plasma was collected to assess lipid peroxidation. Endothelial cells isolated from aorta were evaluated by flow cytometry and fluorescence intensity (FI) emitted by NO-sensitive dye [4,5-diaminofluoresceindiacetate (DAF-2DA)] and by ROS-sensitive dye [dihydroethidium (DHE)]. Vascular reactivity was made by concentration-response curves of acetylcholine. MSG showed hypertension compared with CTR. Treatment with L-NAME increased MAP only in CTR. The MSG induced an increase in the low-frequency (LF) band and a decrease in the high-frequency band of pulse interval. L-NAME treatment increased the LF band of systolic arterial pressure only in CTR without changes in MSG. Lipid peroxidation levels were higher in MSG and were attenuated after L-NAME. In endothelial cells, basal FI to DAF was higher in CTR than in MSG. In both groups, acetylcholine increased FI for DAF from basal. The FI baseline to DHE was higher in MSG than in CTR. Acetylcholine increased FI to DHE in the CTR group, but decreased in MSG animals. We suggest that reduced NO production and increased production of ROS may contribute to hypertension in obese MSG animals.

Sodium nitroprusside activates potassium channels in the vena cava in normotensive but not in hypertensive rats.

Despite the importance of the venous system in the regulation of blood pressure, there are few studies that evaluate venous function in health and disease, and the effects of drugs on venous function. Blood pressure depends directly on the peripheral resistance and cardiac output. Unlike the peripheral resistance, in which the contractile activity of the arteries is the key factor, cardiac output depends primarily on the venomotor tone. An increase in cardiac blood pressure can be caused by an increase in blood volume, structural changes in the walls of the veins, leading to a reduced compliance thereof, or an increase in the contractile activity of venous smooth muscle. This study examined the effect of sodium nitroprusside (SNP), a classical nitric oxide donor, on vascular relaxation in the vena cava from normotensive (2K) and renal hypertensive (2K-1C) rats. We studied the effect of this compound in vena cava rings from normotensive and renal hypertensive rats. We showed for the first time that the vascular relaxation induced by SNP is impaired in vena cavas from hypertensive rats because of an impaired functional activity of potassium channels. Another relevant finding of this study is that the sarcoplasmic reticulum Ca(2+) ATPase is not involved in the venorelaxation induced by SNP.

Deficient regulatory T cell activity and low frequency of IL-17-producing T cells correlate with the extent of cardiomyopathy in human Chagas' disease.

BACKGROUND: Myocardium damage during Chagas' disease results from the immunological imbalance between pro- and production of anti-inflammatory cytokines and has been explained based on the Th1-Th2 dichotomy and regulatory T cell activity. Recently, we demonstrated that IL-17 produced during experimental T. cruzi infection regulates Th1 cells differentiation and parasite induced myocarditis. Here, we investigated the role of IL-17 and regulatory T cell during human Chagas' disease. METHODOLOGY/PRINCIPAL FINDINGS: First, we observed CD4(+)IL-17(+) T cells in culture of peripheral blood mononuclear cells (PBMC) from Chagas' disease patients and we evaluated Th1, Th2, Th17 cytokine profile production in the PBMC cells from Chagas' disease patients (cardiomyopathy-free, and with mild, moderate or severe cardiomyopathy) cultured with T. cruzi antigen. Cultures of PBMC from patients with moderate and severe cardiomyopathy produced high levels of TNF-α, IFN-γ and low levels of IL-10, when compared to mild cardiomyopathy or cardiomyopathy-free patients. Flow cytometry analysis showed higher CD4(+)IL-17(+) cells in PBMC cultured from patients without or with mild cardiomyopathy, in comparison to patients with moderate or severe cardiomyopathy. We then analyzed the presence and function of regulatory T cells in all patients. All groups of Chagas' disease patients presented the same frequency of CD4(+)CD25(+) regulatory T cells. However, CD4(+)CD25(+) T cells from patients with mild cardiomyopathy or cardiomyopathy-free showed higher suppressive activity than those with moderate and severe cardiomyopathy. IFN-γ levels during chronic Chagas' disease are inversely correlated to the LVEF (P = 0.007, r = -0.614), while regulatory T cell activity is directly correlated with LVEF (P = 0.022, r = 0.500). CONCLUSION/SIGNIFICANCE: These results indicate that reduced production of the cytokines IL-10 and IL-17 in association with high levels of IFN-γ and TNF-α is correlated with the severity of the Chagas' disease cardiomyopathy, and the immunological imbalance observed may be causally related with deficient suppressor activity of regulatory T cells that controls myocardial inflammation.