Sodium Nitrite Attenuates Reduced Activity of Vascular Matrix Metalloproteinase-2 and Vascular Hyper-Reactivity and Increased Systolic Blood Pressure Induced by the Placental Ischemia Model of Preeclampsia in Anesthetized Rats.

Preeclampsia is a maternal hypertension disorder associated with vascular dysfunction and fetal and placental growth restrictions. Placental ischemia is suggested as the primary trigger of preeclampsia-associated impairments of both endothelium-derived nitric oxide (NO) and the vascular activity of extracellular matrix metalloproteinase-2 (MMP-2). Reduced uteroplacental perfusion pressure (RUPP) is a placental ischemia model of preeclampsia. Reduction of sodium nitrite to NO may occur during ischemic conditions. However, sodium nitrite effects in the RUPP model of preeclampsia have not yet been investigated. Pregnant rats were divided into four groups: normotensive pregnant rats (Norm-Preg), pregnant rats treated with sodium nitrite (Preg + Nitrite), preeclamptic rats (RUPP), and preeclamptic rats treated with sodium nitrite (RUPP + Nitrite). Maternal blood pressure and fetal and placental parameters were recorded. Vascular function, circulating NO metabolites, and the gelatinolytic activity of vascular MMP-2 were also examined. Sodium nitrite attenuates increased blood pressure, prevents fetal and placental weight loss, counteracts vascular hyper-reactivity, and partially restores NO metabolites and MMP-2 activity. In conclusion, sodium nitrite reduction to NO may occur during RUPP-induced placental ischemia, thereby attenuating increased blood pressure, fetal and placental growth restriction, and vascular hyper-reactivity associated with preeclampsia and possibly restoring NO and MMP-2 activity, which underlie the blood pressure-lowering effects.

Circulating levels of hydrogen sulphide negatively correlate to nitrite levels in gestational hypertensive and preeclamptic pregnant women.

Endothelial dysfunction is a hallmark of preeclampsia and the role of nitric oxide (NO) has been extensively studied in this pregnancy complication. In recent years, hydrogen sulphide (H2 S) has arisen as a new gasotransmitter with an impact on endothelial function. However, the involvement of H2 S in the pathophysiology of preeclampsia is not fully understood, and only a few studies with limited sample size have investigated circulating levels of H2 S in preeclamptic patients. Moreover, H2 S levels have not been previously evaluated in gestational hypertension. Furthermore, the relationship between H2 S and NO in these hypertensive disorders of pregnancy has yet to be determined. We measured H2 S levels in plasma of 120 healthy pregnant women, 88 gestational hypertensive and 62 preeclamptic women. We also measured plasma nitrite in a subset of patients and carried out correlation analysis between plasma H2 S and nitrite in these three groups. We found that plasma H2 S was elevated in preeclampsia and further increased in gestational hypertension compared to healthy pregnancy. Plasma nitrite was reduced in gestational hypertension and preeclampsia, and these levels were negatively correlated with H2 S in both gestational hypertension and preeclampsia, but not in healthy pregnancy. Our results indicate that increases in H2 S may represent a mechanism triggered as an attempt to compensate reduced NO in gestational hypertension and preeclampsia. Future studies are warranted to investigate the mechanisms underlying H2 S/NO interaction on mediating endothelial dysfunction in these hypertensive disorders of pregnancy.

Anticontractile Effect of Perivascular Adipose Tissue But Not of Endothelium Is Enhanced by Hydrogen Sulfide Stimulation in Hypertensive Pregnant Rat Aortae.

Perivascular adipose tissue (PVAT) modulates the vascular tone. Hydrogen sulfide (H2S) is synthetized by cystathionine gamma-lyase (CSE) in brown PVAT. Modulation of vascular contractility by H2S is, in part, adenosine triphosphate (ATP)-sensitive potassium channels dependent. However, the role of PVAT-derived H2S in hypertensive pregnancy (HTN-Preg) is unclear. Therefore, we aimed to examine the involvement of H2S in the anticontractile effect of PVAT in aortae from normotensive and hypertensive pregnant rats. To this end, phenylephrine-induced contractions in the presence and absence of PVAT and endothelium in aortae from normotensive pregnant (Norm-Preg) and HTN-Preg rats were investigated. Maternal blood pressure, fetal-placental parameters, angiogenesis-related biomarkers, and H2S levels were also assessed. We found that circulating H2S is elevated in hypertensive pregnancy associated with angiogenic imbalance, fetal and placental growth restrictions, which revealed that there is H2S pathway activation. Moreover, under stimulated H2S formation PVAT, but not endothelium, reduced phenylephrine-induced contractions in aortae from HTN-Preg rats. Also, H2S synthesis inhibitor abolished anticontractile effects of PVAT and endothelium. Furthermore, anticontractile effect of PVAT, but not of endothelium, was eliminated by ATP-sensitive potassium channels blocker. In accordance, increases in H2S levels in PVAT and placenta, but not in aortae without PVAT, were also observed. In conclusion, anticontractile effect of PVAT is lost, at least in part, in HTN-Preg aortae and PVAT effect is ATP-sensitive potassium channels dependent in normotensive and hypertensive pregnant rat aortae. PVAT but not endothelium is responsive to the H2S stimulation in hypertensive pregnant rat aortae, implying a key role for PVAT-derived H2S under endothelial dysfunction.

Impaired Pulmonary Arterial Vasoconstriction and Nitric Oxide-Mediated Relaxation Underlie Severe Pulmonary Hypertension in the Sugen-Hypoxia Rat Model.

Pulmonary vasoreactivity could determine the responsiveness to vasodilators and, in turn, the prognosis of pulmonary hypertension (PH). We hypothesized that pulmonary vasoreactivity is impaired, and we examined the underlying mechanisms in the Sugen-hypoxia rat model of severe PH. Male Sprague-Dawley rats were injected with Sugen (20 mg/kg s.c.) and exposed to hypoxia (9% O2) for 3 weeks, followed by 4 weeks in normoxia (Su/Hx), or treated with Sugen alone (Su) or hypoxia alone (Hx) or neither (Nx). After hemodynamic measurements, the heart was assessed for right ventricular hypertrophy (Fulton's index); the pulmonary artery, aorta, and mesenteric arteries were isolated for vascular function studies; and contractile markers were measured in pulmonary arteries using quantitative polymerase chain reaction (PCR). Other rats were used for morphometric analysis of pulmonary vascular remodeling. Right ventricular systolic pressure and Fulton's index were higher in Su/Hx versus Su, Hx, and Nx rats. Pulmonary vascular remodeling was more prominent in Su/Hx versus Nx rats. In pulmonary artery rings, contraction to high KCl (96 mM) was less in Su/Hx versus Nx and Su, and phenylephrine-induced contraction was reduced in Su/Hx versus Nx, Hx, and Su. Acetylcholine (ACh)-induced relaxation was less in Su/Hx versus Nx and Hx, suggesting reduced endothelium-dependent vasodilation. ACh relaxation was inhibited by nitric oxide synthase (NOS) and guanylate cyclase blockade in all groups, suggesting a role of the NO-cGMP pathway. Nitrate/nitrite production in response to ACh was less in Su/Hx versus Nx, supporting reduced endothelial NO production. Sodium nitroprusside (10-8 M) caused less relaxation in Su/Hx versus Nx, Hx, and Su, suggesting a decreased responsiveness of vascular smooth muscle (VSM) to vasodilators. Neither contraction nor relaxation differed in the aorta or mesenteric arteries of all groups. PCR analysis showed decreased expression of contractile markers in pulmonary artery of Su/Hx versus Nx. The reduced responsiveness to vasoconstrictors and NO-mediated vasodilation in the pulmonary, but not systemic, vessels may be an underlying mechanism of severe PH in Su/Hx rats and appears to involve attenuation of the NO relaxation pathway and a switch of pulmonary VSM cells to a synthetic less reactive phenotype.

Increases in placental nitric oxide, but not nitric oxide-mediated relaxation, underlie the improvement in placental efficiency and antihypertensive effects of hydrogen sulphide donor in hypertensive pregnancy.

Dysregulation of hydrogen sulphide (H2 S) producing enzymes has been related to hypertensive pregnancy, and H2 S donor, sodium hydrosulphide (NaHS) exerts antihypertensive effects, modulates angiogenic factors production and acts as an antioxidant. Moreover, reduction in nitric oxide (NO) bioavailability is related to hypertensive pregnancy and H2 S may interact with NO, modulating its production. We aimed to investigate the NaHS effects in hypertension-in-pregnancy and also in feto-placental parameters. Female Wistar rats (200-250 g) were mated and desoxycorticosterone acetate injections followed by replacement of water by 0.9% saline solution were used to induce hypertensive pregnancy. Rats were divided into four groups: normal pregnant (Norm-Preg), pregnant + NaHS (Preg+NaHS), hypertensive pregnant (HTN-Preg) and HTN-Preg+NaHS. Systolic blood pressure was increased in HTN-Preg and this increase was blunted in HTN-Preg+NaHS. Fetal and placental weights were decreased in HTN-Preg animals, while fetal growth restriction was improved in HTN-Preg+NaHS. Placental weight was lower in HTN-Preg+NaHS than in HTN-Preg; however, placental efficiency was re-established in HTN-Preg+NaHS rats. We observed that a partial contribution of placental NO, but not changes in anti-angiogenic factors may mediate the increases in placental efficiency in HTN-Preg+NaHS. HTN-Preg presented thoracic aorta hyperreactivity to phenylephrine while NaHS treatment blunted this hyperreactivity, which seems not to be related to NO-mediated relaxation induced by acetylcholine. Therefore, changes in vascular responsiveness promoted by NaHS treatment may underlie the beneficial effects in systolic blood pressure and feto-placental parameters in our study.

Sodium nitrite attenuates hypertension-in-pregnancy and blunts increases in soluble fms-like tyrosine kinase-1 and in vascular endothelial growth factor.

Preeclampsia is a pregnancy-associated disorder characterized by hypertension with uncertain pathogenesis. Increases in antiangiogenic soluble fms-like tyrosine kinase-1 (sFlt-1) and reductions in nitric oxide (NO) bioavailability have been observed in preeclamptic women. However, the specific mechanisms linking these detrimental changes to the hypertension-in-pregnancy are not clearly understood. In this regard, while recent findings have suggested that nitrite-derived NO formation exerts antihypertensive and antioxidant effects, no previous study has examined these responses to orally administered nitrite in hypertension-in-pregnancy. We then hypothesized restoring NO bioavailability with sodium nitrite in pregnant rats upon NO synthesis inhibition with N(omega)-nitro-l-arginine methyl ester (L-NAME) attenuates hypertension and high circulating levels of sFlt-1. Number and weight of pups and placentae were recorded to assess maternal-fetal interface. Plasma sFlt-1, vascular endothelial growth factor (VEGF) and biochemical determinants of NO formation and of antioxidant function were measured. We found that sodium nitrite blunts the hypertension-in-pregnancy and restores the NO bioavailability, and concomitantly prevents the L-NAME-induced high circulating sFlt-1 and VEGF levels. Also, our results suggest that nitrite-derived NO protected against reductions in litter size and placental weight caused by L-NAME, improving number of viable and resorbed fetuses and antioxidant function. Therefore, the present findings are consistent with the hypothesis that nitrite-derived NO may possibly be the driving force behind the maternal and fetal beneficial effects observed with sodium nitrite during hypertension-in-pregnancy. Certainly further investigations are required in preeclampsia, since counteracting the damages to the mother and fetal sides resulting from hypertension and elevated sFlt-1 levels may provide a great benefit in this gestational hypertensive disease.

Maternal pregnancy hypertension impairs nitric oxide formation and results in increased arterial blood pressure in first-generation offspring female rats.

OBJECTIVES: Maternal endothelial dysfunction in pregnancy hypertension is related to impairment of nitric oxide (NO) formation. However, NO levels and hemodynamic repercussions on the female offspring remain unclear. Therefore, this study hypothesized that maternal pregnancy hypertension reduces circulating NO metabolites and increases arterial blood pressure in first-generation offspring female rats. STUDY DESIGN: Descendant female rats were distributed in four groups as follows: virgin offspring of normotensive (VN) and hypertensive (VH) mothers and pregnant offspring of normotensive (PN) and hypertensive (PH) mothers. Hemodynamic and biochemical analyses were performed. MAIN OUTCOME MEASURES: The systolic (SBP) and diastolic (DBP) blood pressure, heart rate (HR), and body weight were measured. NO metabolites in plasma, NO formation in human umbilical vein endothelial cells (HUVECs) incubated with plasma, and endothelial NO synthase (eNOS) expression in aortas were determined. RESULTS: Increased SBP, DBP, and reduced HR were found on the 60 days of life in the VH group, whereas the PH group showed increased SBP and HR on pregnancy day 7. All groups showed no differences in body weight gain and eNOS expression. Plasma levels of NO metabolites were increased in the PN compared to the other groups. Increases in the NO formation were greater in HUVECs incubated with plasma from VN and PN groups compared to the VH and PH groups. CONCLUSIONS: Female virgin and pregnant first-generation offspring rats from hypertensive pregnant mothers may have negative cardiovascular repercussions featured by increases in SBP, and possibly impaired NO formation is involved.

Isoflurane increases the activity of the vascular matrix metalloproteinase-2 in non-pregnant rats and increases the nitric oxide metabolites in pregnancy.

Surgeries that require general anesthesia occur in 1.5-2% of gestations. Isoflurane is frequently used because of its lower possibility of affecting fetal growth. Therefore, we examined the isoflurane anesthesia-induced effects on maternal hemodynamic and vascular changes. We hypothesized that isoflurane would enhance endothelium-dependent vasodilation as a consequence of increased nitric oxide and decreased metalloproteinases (MMPs). Female rats (n=28) were randomized into 4 groups (7 rats/group): conscious (non-anesthetized) non-pregnant group, non-pregnant anesthetized group, conscious pregnant group, and pregnant anesthetized group. Anesthesia was performed on the 20th pregnancy day, and hemodynamic parameters were monitored. Nitric oxide metabolites, gelatinolytic activity of MMP-2 and MMP-9, and the vascular function were assessed. Isoflurane caused no significant hemodynamic changes in pregnant compared with non-pregnant anesthetized group. Impaired acetylcholine-induced relaxations were observed only in conscious non-pregnant group (by approximately 62%) versus 81% for other groups. Phenylephrine-induced contractions were greater in endothelium-removed aorta segments of both pregnant groups (with or without isoflurane) compared with non-pregnant groups. Higher nitric oxide metabolites were observed in anesthetized pregnant in comparison with the other groups. Reductions in the 75 kDa activity and concomitant increases in 64 kDa MMP-2 isoforms were observed in aortas of pregnant anesthetized (or not) groups compared with conscious non-pregnant group. Isoflurane anesthesia shows stable effects on hemodynamic parameters and normal MMP-2 activation in pregnancy. Furthermore, there were increases in nitric oxide bioavailability, suggesting that isoflurane provides protective actions to the endothelium in pregnancy.

Elevated plasma hemoglobin levels increase nitric oxide consumption in experimental and clinical acute pulmonary thromboembolism.

OBJECTIVES: We examined whether experimental lung embolization with autologous blood clots or with the infusion of microspheres increase cell-free hemoglobin levels and nitric oxide consumption by plasma samples from anesthetized lambs. These parameters were also measured in patients with acute pulmonary thromboembolism at baseline conditions and after thrombolysis, and in healthy controls. DESIGN: Controlled animal and clinical studies. SETTING: University research laboratory and university hospital. SUBJECTS: Sheep and humans. INTERVENTIONS: Anesthetized lambs were embolized with intravenous injections of autologous blood clots or repeated injections of 300 µm microspheres. Control animals received saline. Blood samples were drawn from patients with acute pulmonary thromboembolism at baseline conditions and after thrombolytic therapy with streptokinase or alteplase. MEASUREMENTS AND MAIN RESULTS: Hemodynamic measurements were performed and plasma cell-free hemoglobin concentrations were measured. A nitric oxide consumption assay was used to measure nitric oxide consumption by plasma samples. Embolization with blood clots or microspheres increased mean pulmonary artery pressure from ~15 to ~40 mm Hg in lambs. Both plasma hemoglobin concentrations and nitric oxide consumption increased in proportion to the hemodynamic alterations and correlated significantly. Patients with acute pulmonary thromboembolism had higher plasma hemoglobin concentrations and nitric oxide consumption than healthy controls. Thrombolysis with streptokinase or alteplase further increased both parameters, which peaked 1-3 days after thrombolysis. CONCLUSIONS: Our results show consistent evidence indicating a new mechanism involving increased hemoglobin decompartmentalization and augmented nitric oxide consumption, possibly contributing to the hemodynamic derangement of acute pulmonary thromboembolism.

Vasodilator Responses of Perivascular Adipose Tissue-Derived Hydrogen Sulfide Stimulated with L-Cysteine in Pregnancy Hypertension-Induced Endothelial Dysfunction in Rats.

Endothelium-derived nitric oxide (NO)-induced vasodilation is impaired in pregnancy hypertension. However, the role of perivascular adipose tissue (PVAT)-derived hydrogen sulfide (H2S), as an alternative for counteracting vascular dysfunction, is incompletely clear in hypertensive disorders of pregnancy. Therefore, PVAT-derived H2S-induced vasodilation was investigated in pregnancy hypertension-induced endothelial dysfunction. Non-pregnant (Non-Preg) and pregnant (Preg) rats were submitted (or not) to the deoxycorticosterone (DOCA)-salt protocol and assigned as follows (n = 10/group): Non-Preg, Non-Preg+DOCA, Preg, and Preg+DOCA groups. Systolic blood pressure (SBP), angiogenesis-related factors, determinant levels of H2S (PbS), NO (NOx), and oxidative stress (MDA) were assessed. Vascular changes were recorded in thoracic aortas with PVAT and endothelium (intact and removed layers). Vasorelaxation responses to the substrate (L-cysteine) for the H2S-producing enzyme cystathionine-γ-lyase (CSE) were examined in the absence and presence of CSE-inhibitor DL-propargylglycine (PAG) in thoracic aorta rings pre-incubated with cofactor for CSE (pyridoxal-5 phosphate: PLP) and pre-contracted with phenylephrine. Hypertension was only found in the Preg+DOCA group. Preg+DOCA rats showed angiogenic imbalances and increased levels of MDA. PbS, but not NOx, showed increased levels in the Preg+DOCA group. Pre-incubation with PLP and L-cysteine elevated determinants of H2S in PVAT and placentas of Preg-DOCA rats, whereas no changes were found in the aortas without PVAT. Aortas of Preg-DOCA rats showed that PVAT-derived H2S-dependent vasodilation was greater compared to endothelium-derived H2S, whereas PAG blocked these responses. PVAT-derived H2S endogenously stimulated with the amino acid L-cysteine may be an alternative to induce vasorelaxation in endothelial dysfunction related to pregnancy hypertension.

Pravastatin Prevents Increases in Activity of Metalloproteinase-2 and Oxidative Stress, and Enhances Endothelium-Derived Nitric Oxide-Dependent Vasodilation in Gestational Hypertension.

Pre-eclampsia (PE) is a hypertensive disorder of pregnancy and has been associated with placental growth restriction. The pre-eclamptic placenta releases free radicals to maternal circulation, thus increasing oxidative stress. An impaired redox state leads to reduction in circulating nitric oxide (NO) levels and activation of extracellular matrix metalloproteinases (MMPs). However, activation of MMPs induced by oxidative stress is still unclear in PE. Antioxidant effects have been demonstrated with the use of pravastatin. Therefore, we hypothesized that pravastatin protects against oxidative stress-induced activation of MMPs in a rat model of PE. The animals were divided into four groups: normotensive pregnant rats (Norm-Preg); pregnant rats treated with pravastatin (Norm-Preg + Prava); hypertensive pregnant rats (HTN-Preg); and hypertensive pregnant rats treated with pravastatin (HTN-Preg + Prava). The deoxycorticosterone acetate (DOCA) and sodium chloride (DOCA-salt) model was used to induce hypertension in pregnancy. Blood pressure, and fetal and placental parameters were recorded. The gelatinolytic activity of MMPs, NO metabolites and lipid peroxide levels were also determined. Endothelium function was also examined. Pravastatin attenuated maternal hypertension, prevented placental weight loss, increased NO metabolites, inhibited increases in lipid peroxide levels, and reduced the activity of MMP-2, and these effects were observed along with enhanced endothelium-derived NO-dependent vasodilation. The present results provide evidence that pravastatin protects against activation of MMP-2 induced by oxidative stress in pre-eclamptic rats. These findings may also involve improvement in endothelial function related to NO and antihypertensive effects of pravastatin, thus suggesting pravastatin as a therapeutic intervention for PE.

Pregnancy hypertension-associated endothelial dysfunction is attenuated by isoflurane anesthesia: Evidence of protective effect related to increases in nitric oxide.

AIMS: Pregnancy hypertension-induced endothelial dysfunction associated with impairment of nitric oxide (NO) bioavailability and hemodynamic derangements is a challenging for urgent procedures requiring maternal anesthesia. The volatile anesthetic isoflurane has demonstrated NO-associated protective effects. However, this isoflurane-induced effect is still unclear in pregnancy hypertension. Therefore, the present study examined the potential protective effects of isoflurane anesthesia on endothelial dysfunction and hemodynamic changes induced by hypertensive pregnancy associated with fetal and placental growth restrictions. MATERIALS AND METHODS: Animals were distributed into four groups: normotensive pregnant rats (Preg), anesthetized pregnant rats (Preg+Iso), hypertensive pregnant rats (HTN-Preg), and anesthetized hypertensive pregnant rats (HTN-Preg+Iso). Systolic and diastolic pressures, mean arterial pressure (MAP), heart rate, fetal and placental weights, vascular contraction, endothelium-derived NO-dependent vasodilation, and NO levels were assessed. The vascular endothelial growth factor (VEGF) levels and endothelial NO synthase (eNOS) Serine (1177) phosphorylation (p-eNOS) expression were also examined. KEY FINDINGS: Isoflurane produced more expressive hypotensive effects in the HTN-Preg+Iso versus Preg+Iso group, with respective reductions in MAP by 50 ± 13 versus 25 ± 4 mmHg (P < 0.05). Also, HTN-Preg+Iso compared to the HTN-Preg group showed (respectively) preventions against the weight loss of the fetuses (4.0 ± 0.6 versus 2.8 ± 0.6 g, P < 0.05) and placentas (0.37 ± 0.06 versus 0.30 ± 0.06 mg, P < 0.05), hyper-reactive vasocontraction response (1.8 ± 0.4 versus 2.8 ± 0.6 g, P < 0.05), impaired endothelium-derived NO-dependent vasodilation (84 ± 8 versus 50 ± 17 %, P < 0.05), reduced VEGF levels (147 ± 46 versus 25 ± 13 pg/mL, P < 0.05), and decreased p-eNOS expression (0.24 ± 0.07 versus 0.09 ± 0.05 arbitrary units, P < 0.05). SIGNIFICANCE: Isoflurane anesthesia protects maternal endothelial function in pregnancy hypertension, and possibly endothelium-derived NO is involved.

Sevoflurane and isoflurane anesthesia induce redox imbalance, but only sevoflurane impairs vascular contraction.

Volatile anesthetics may cause vascular dysfunction; however, underlying effects are unclear. The aim of the present study was to investigate whether sevoflurane and isoflurane affect vascular function, nitric oxide (NO) bioavailability, and biomarkers of oxidative stress and inflammation. Wistar rats were divided into three experimental groups: Not anesthetized (control group) or submitted to anesthesia with isoflurane (Iso group) or sevoflurane (Sevo group). Hemodynamic parameters were monitored during anesthesia, and blood gas values and biochemical determinants were analyzed. Isometric contractions were recorded in aortic rings. Vasoconstriction induced by potassium chloride (KCl) and phenylephrine (Phe) were measured. No differences in hemodynamic parameters and blood gasses variables were observed. Impaired KCl and Phe-induced contractions were observed in endothelium-intact aorta of Sevo compared to Iso and Control groups. Redox imbalance was found in Sevo and Iso groups. Reduced NO bioavailability and increased activity of matrix metalloproteinase 2 (MMP-2) were observed in Sevo, but not in the Iso group. While reduced IL-10 and IL-1ß were observed in Sevo, increases in IL-1ß in the Iso group were found. Sevoflurane, but not isoflurane, anesthesia impairs vasocontraction, and reduced NO and cytokines and increased MMP-2 activity may be involved in vascular dysfunction after sevoflurane anesthesia.

Sildenafil attenuates oxidative stress and endothelial dysfunction in lead-induced hypertension.

Lead (Pb) reduces NO bioavailability, impairs the antioxidant system, and increases the generation of reactive oxygen species (ROS). Pb-induced oxidative stress may be responsible for the associated endothelial dysfunction. Sildenafil has shown nitric oxide (NO)-independent action, including antioxidant effects. Therefore, we examined the effects of sildenafil on oxidative stress, reductions of NO and endothelial dysfunction in Pb-induced hypertension. Wistar rats were distributed into three groups: Pb, Pb + sildenafil and Sham. Blood pressure and endothelium-dependent vascular function were recorded. We also examined biochemical determinants of lipid peroxidation and antioxidant function. ROS levels, NO metabolites and NO levels in human umbilical vein endothelial cells (HUVECs) were also evaluated. Sildenafil prevents impairment of endothelium-dependent NO-mediated vasodilation and attenuates Pb-induced hypertension, reduces ROS formation, enhances superoxide dismutase (SOD) activity and antioxidant capacity in plasma and increases NO metabolites in plasma and HUVECs culture supernatants, while no changes were found on measurement of NO released from HUVECs incubated with plasma of the Pb and Pb + sildenafil groups compared with the sham group. In conclusion, sildenafil protects against ROS-mediated inactivation of NO, thus preventing endothelial dysfunction and attenuating Pb-induced hypertension, possibly through antioxidant effects.

A new look at the role of nitric oxide in preeclampsia: Protein S-nitrosylation.

The formation of S-nitrosothiols (SNOs) occurs with the reaction of nitric oxide (NO) and free thiol groups in proteins. This process, called S-nitrosylation, allows NO to interfere with or even modulate a variety of cellular functions, culminating with the modification of protein trafficking, redox state, and cell cycle. Furthermore, NO plays a role in modulating a wide range of functions in endothelial cells specifically, including inflammation, apoptosis, permeability, migration, and cell growth. As such, NO acts as a mediator in several physiological processes. The interaction between endothelial nitric oxide synthase (eNOS) and proteins that are to be targeted for S-nitrosylation is a key determinant of the specificity of NO signaling. Deficits in the bioavailability of NO have been associated with pregnancy-related disorders, such as preeclampsia (PE). The study of S-nitrosylation in PE, as well as the identification of targeted proteins, may contribute to a better understanding of its pathophysiology and the development of drugs for the treatment of PE patients. In this review, we aimed to present the mechanism of S-nitrosylation, the regulatory pathways, and some proteins by which S-nitrosylation can modulate NO availability with a potential impact on PE.

Metalloproteinase inhibition protects against cardiomyocyte injury during experimental acute pulmonary thromboembolism.

OBJECTIVES: Up-regulated matrix metalloproteinases may be involved in the development of cardiomyocyte injury and the degradation of troponin associated with acute pulmonary thromboembolism. We examined whether pretreatment with doxycycline (a nonspecific matrix metalloproteinase inhibitor) protects against cardiomyocyte injury associated with acute pulmonary thromboembolism. DESIGN: Controlled animal study. SETTING: University research laboratory. SUBJECTS: Mongrel dogs. INTERVENTIONS: Anesthetized animals received doxycycline (10 mg/kg intravenously) or saline and acute pulmonary thromboembolism was induced with autologous blood clots injected into the right atrium. Control animals received doxycycline (or saline). MEASUREMENTS AND MAIN RESULTS: Hemodynamic measurements were performed, and acute pulmonary thromboembolism increased baseline mean pulmonary arterial pressure and pulmonary vascular resistance by approximately 160% and 362%, respectively (both p < .05), 120 mins after acute pulmonary thromboembolism. Pretreatment with doxycycline attenuated these increases (to 125% and 232%, respectively; both p < .05). Although acute pulmonary thromboembolism tended to increase the right ventricle maximum rate of isovolumic pressure development and the maximum rate of isovolumic pressure decay, doxycycline produced no effects on these parameters. Gelatin zymograms of right ventricle showed that acute pulmonary thromboembolism marginally increased matrix metalloproteinase-9 (but not matrix metalloproteinase-2) levels in the right ventricle. A fluorometric assay to assess net matrix metalloproteinase activities showed that acute pulmonary thromboembolism increased matrix metalloproteinase activities in the right ventricle by >100% (p < .05), and this finding was confirmed by in situ zymography of the right ventricle. Doxycycline attenuated acute pulmonary thromboembolism-induced increases in right ventricle matrix metalloproteinase activities. Acute pulmonary thromboembolism induced neutrophil accumulation in the right ventricle, as estimated by myeloperoxidase activity, and doxycycline blunted this effect (p < .05). Serum cardiac troponin I concentrations, which reflect cardiomyocyte injury, increased after acute pulmonary thromboembolism, and this increase was attenuated by pretreatment with doxycycline (p < .05). CONCLUSIONS: We found evidence supporting the idea that acute pulmonary thromboembolism is associated with increased matrix metalloproteinase activities in the right ventricle, which may lead to degradation of sarcomeric proteins, including cardiac troponin I. Inhibition of matrix metalloproteinases may be an effective therapeutic intervention in the management of acute pulmonary thromboembolism.

Hemodynamic effects of inducible nitric oxide synthase inhibition combined with sildenafil during acute pulmonary embolism.

While endogenous nitric oxide (NO) may be relevant to the beneficial hemodynamic effects produced by sildenafil during acute pulmonary embolism (APE), huge amounts of inducible NO synthase (iNOS)-derived NO may contribute to lung injury. We hypothesized that iNOS inhibition with S-methylisothiourea could attenuate APE-induced increases in oxidative stress and pulmonary hypertension and, therefore, could improve the beneficial hemodynamic and antioxidant effects produced by sildenafil during APE. Hemodynamic evaluations were performed in non-embolized dogs treated with saline (n=4), S-methylisothiourea (0.01 mg/kg followed by 0.5 mg/kg/h, n=4), sildenafil (0.3 mg/kg, n=4), or S-methylisothiourea followed by sildenafil (n=4), and in dogs that received the same drugs and were embolized with silicon microspheres (n=8 for each group). Plasma nitrite/nitrate (NOx) and thiobarbituric acid reactive substances (TBARS) concentrations were determined by Griess and a fluorometric assay, respectively. APE increased mean pulmonary arterial pressure (MPAP) and pulmonary vascular resistance index (PVRI) by 25±1.7 mm Hg and by 941±34 dyn s cm(-5) m(-2), respectively. S-methylisothiourea neither attenuated APE-induced pulmonary hypertension, nor enhanced the beneficial hemodynamic effects produced by sildenafil after APE (>50% reduction in pulmonary vascular resistance). While sildenafil produced no change in plasma NOx concentrations, S-methylisothiourea alone or combined with sildenafil blunted APE-induced increases in NOx concentrations. Both drugs, either alone or combined, produced antioxidant effects. In conclusion, although iNOS-derived NO may play a key role in APE-induced oxidative stress, our results suggest that the iNOS inhibitor S-methylisothiourea neither attenuates APE-induced pulmonary hypertension, nor enhances the beneficial hemodynamic effects produced by sildenafil.

Effects of fast versus slow-releasing hydrogen sulfide donors in hypertension in pregnancy and fetoplacental growth restriction.

Hydrogen sulfide (H2S) is a vasorelaxant gas with therapeutic potential in several diseases. However, effects of H2S donors in hypertensive pregnancy complicated by feto-placental growth restriction are unclear. Therefore, we aimed to examine and compare the effects of fast-releasing H2S donor (sodium hydrosulfide-NaHS) and slow-releasing H2S donor (GYY4137) in hypertension-in-pregnancy. Pregnant rats were distributed into four groups: normal pregnancy (Norm-Preg), hypertensive pregnancy (HTN-Preg), hypertensive pregnancy + NaHS (HTN-Preg + NaHS), and hypertensive pregnancy + GYY4137 (HTN-Preg + GYY). Systolic blood pressure, plasma H2S levels, fetal and placental weights, number of viable fetuses, litter size, and endothelium-dependent vasodilation were examined. Also, oxidative stress was assessed in placenta. We found that GYY4137 attenuated hypertension on gestational days 16 and 18, while NaHS presented antihypertensive effect only on gestational day 18. GYY4137, but not NaHS, increased plasma H2S levels. Greater fetal and placental weights were found with GYY4137 than NaHS treatment. Also, HTN-Preg + NaHS presented further reductions in placental weights when compared to HTN-Preg group. Number of viable fetuses and litter size presented no significant changes. GYY4137 reduced placental oxidative stress caused by hypertension, while greater increases in oxidative stress were found in HTN-Preg + NaHS than HTN-Preg group. Hypertensive pregnancy caused impaired endothelium-dependent vasodilation, while GYY4137 and NaHS treatments blunted endothelial dysfunction. Endothelium-dependent vasodilation was completely blocked by the nitric oxide synthase inhibitor. We conclude that slow-releasing H2S donor GYY4137 is advantageous compared with fast-releasing H2S-donor NaHS to attenuate hypertension-in-pregnancy and to protect against feto-placental growth restriction and oxidative stress.

Placental nitric oxide formation and endothelium-dependent vasodilation underlie pravastatin effects against angiogenic imbalance, hypertension in pregnancy and intrauterine growth restriction.

Pre-eclampsia and hypertensive disorders of pregnancy are frequently associated with foeto-placental growth restriction, and that may be triggered by angiogenic imbalance and endothelial dysfunction. Impaired nitric oxide (NO) bioavailability seems to be involved in these pathophysiological changes observed in hypertensive pregnancy. Pravastatin has shown efficacy and to be safe during hypertension in pregnancy. However, NO involvement in pravastatin effects during maternal hypertension and foeto-placental development is unclear. Therefore, we aimed to examine pravastatin effects on placental NO formation, endothelium-dependent vasodilation, systolic blood pressure and foeto-placental development in hypertensive pregnant rats. Biochemical determinants of angiogenesis and oxidative stress were also assessed. Pregnant rats were distributed into four groups: normal pregnancy (Norm-Preg), pregnancy+pravastatin (Preg-Prava), hypertensive pregnancy (HTN-Preg) and hypertensive pregnancy+pravastatin (HTN-Preg+Prava). Our results showed that pravastatin treatment blunts hypertension and foeto-placental growth restriction. Also, increases in placental NO levels were found in the HTN-Preg+Prava group. Pravastatin prevents impaired endothelium-dependent acetylcholine-induced vasodilation, exacerbated contractile response to phenylephrine and increases in oxidative stress in the HTN-Preg+Prava group. Increased soluble fms-like tyrosine kinase-1-to-placental growth factor (sFlt-1/PlGF) ratio is reversed by pravastatin treatment in the HTN-Preg+Prava group. We conclude that NO formation and endothelium-dependent vasodilation underlie pleiotropic effects associated with pravastatin treatment against hypertension in pregnancy, intrauterine growth restriction, vascular dysfunction and angiogenic imbalance.

Reductions of Circulating Nitric Oxide are Followed by Hypertension during Pregnancy and Increased Activity of Matrix Metalloproteinases-2 and -9 in Rats.

Hypertensive pregnancy has been associated with reduced nitric oxide (NO), bioavailability, and increased activity of matrix metalloproteinases (MMPs). However, it is unclear if MMPs activation is regulated by NO during pregnancy. To this end, we examined activity of MMP-2 and MMP-9 in plasma, placenta, uterus and aorta, NO bioavailability, oxidative stress, systolic blood pressure (SBP), and fetal-placental development at the early, middle, and late pregnancy stages in normotensive and Nω-Nitro-L-arginine methyl-ester (L-NAME)-induced hypertensive pregnancy in rats. Reduced MMP-2 activity in uterus, placenta, and aorta and reduced MMP-9 activity in plasma and placenta with concomitant increased NO levels were found in normotensive pregnant rats. By contrast, increased MMP-2 activity in uterus, placenta, and aorta, and increased MMP-9 activity in plasma and placenta with concomitant reduced NO levels were observed in hypertensive pregnant rats. Also, elevated oxidative stress was displayed by hypertensive pregnant rats at the middle and late stages. These findings in the L-NAME-treated pregnant rats were also followed by increases in SBP and associated with fetal growth restrictions at the middle and late pregnancy stages. We concluded that NO bioavailability may regulate MMPs activation during normal and hypertensive pregnancy.