Relaxation Of Aorta Research Articles

Effects of a novel selenium substituted-sugar (1,4-anhydro-4-seleno-d-talitol, SeTal) on human coronary artery cell lines and mouse aortic rings

Chronic low-grade inflammation and oxidative damage are strongly associated with pathologies including cardiovascular disease. As a consequence, there is considerable interest in agents that mitigate damage. Selenium compounds can act as potent protective agents against oxidation due to the high reactivity and nucleophilicity of the selenium atom. 1,4-Anhydro-4-seleno-d-talitol (SeTal, a novel water-soluble selenium-based sugar) is a potent oxidant scavenger in vitro and in human plasma. Here we show that SeTal is highly stable in solutions that mimic biological fluids and the gastrointestinal tract, and is not rapidly degraded or metabolized unlike some other selenium-containing compounds. SeTal remains intact during extended storage, and it rapidly penetrates into, and effluxes from, primary human coronary artery endothelial and smooth muscle cells, but does not induce loss of metabolic activity, or modulate cell survival and growth rates at concentrations ≤2 mM. Steady-state intracellular concentrations can reach 2–10 μM. SeTal affords protection against H2O2- and HOCl-mediated oxidative damage, with this being independent of the concentration or activities of the selenium-dependent protective enzymes TrxR and GPx. Protection was observed with both concurrent drug and oxidant administration and also (to a lesser extent) with cellular pre-loading. SeTal also affords protection to isolated arterial segments, with the compound decreasing HOCl (50 μΜ) mediated effects on aortic ring relaxation, consistent with the preservation of NO bioavailability. The stability, bioavailability and protective actions of this compound, suggest that it is worthy of further investigation as a protective agent, particularly in the area of cardiovascular disease.

Metabolite profiling, arginase inhibition and vasorelaxant activity of Cornus mas, Sorbus aucuparia and Viburnum opulus fruit extracts

The present study investigated the effects of Cornus mas, Sorbus aucuparia and Viburnum opulus fruit extracts on arginase activity and arterial vasodilation. V. opulus fruit extract exerted the highest vasorelaxant activity in phenylephrine precontracted rat aortic rings (EC50 = 6.31 ± 1.61 μg/mL) and a significant inhibition of arginase (IC50 = 71.02 ± 3.06 μg/mL). By contrast, S. aucuparia and C. mas fruit extracts showed no important anti-arginase activity and a significantly weaker activity in the rat aortic rings relaxation assay (EC50 = 100.9 ± 11.63 and 78.52 ± 8.59 μg/mL, respectively). For all extracts, the main mechanism of vasodilation was proven to be endothelium-dependent. HPLC-ESI-Q-TOF-MS/MS studies revealed a very complex metabolite profiling in all three extracts with chlorogenic acid accounting for 30.89, 0.72 and 2.03 mg/g in V. opulus, C. mas and S. aucuparia fruit extracts, respectively. All extracts were declared non-toxic in the brine shrimp acute toxicity test. Our study highlights potential benefits of V. opulus fruit extract in diseases associated with endothelial dysfunction and impaired vasodilation.

Influence of type-4 dipeptidyl peptidase inhibition on endothelium-dependent relaxation of aortae from a db/db mouse model of type 2 diabetes: a comparison with the effect of glimepiride.

PurposeThe aim of this study was to investigate the effects of the type-4 dipeptidyl peptidase (DPP-4) inhibitors linagliptin and vildagliptin as well as the sulfonylurea glimepiride on endothelium-dependent relaxation of aortae from female db/db mice with established hyperglycemia to determine whether these treatments were able to attenuate diabetes-induced endothelial dysfunction.Materials and methodsThe mice were treated with glimepiride (2 mg/kg po per day, weeks 1–6, n=12), glimepiride plus vildagliptin (glimepiride 2 mg/kg po per day, weeks 1–6; vildagliptin 3 mg/kg po per day, weeks 4–6, n=11), glimepiride plus linagliptin (glimepiride 2 mg/kg po per day, weeks 1–6; linagliptin 3 mg/kg po per day, weeks 4–6, n=11) or linagliptin (3 mg/kg po per day, weeks 1–6, n=12). Endothelium-dependent relaxation using acetylcholine was assessed in the absence and presence of pharmacological tools (TRAM-34 1 μM; apamin 1 μM; N-nitro-L-arginine [L-NNA] 100 μM; 1H-[1,2,4]oxadiazolo [4,3-a]quinoxalin-1-one [ODQ] 10 μM) to distinguish relaxation mediated by nitric oxide (NO).ResultsLinagliptin was associated with a significant improvement in endothelium-dependent relaxation (ACh Rmax; db/db 41±1%, linagliptin 73±6%, p<0.05). The enhanced response was maintained in the presence of TRAM-34+ apamin (ACh Rmax; db/db 23±6%, linagliptin 60±6%, p<0.01), ie, when the endothelium-dependent relaxation was mediated by NO. There was no evidence for a contribution from KCa channel opening to responses under any conditions. Glimepiride had no effect on endothelium-dependent relaxation when given alone (ACh Rmax 38±3%). The addition of linagliptin or vildagliptin to glimepiride did not significantly improve endothelium-dependent relaxation. All treatments caused some decrease in aortic superoxide production but the effect of linagliptin was significantly greater than glimepiride (linagliptin 534±60 relative luminescence unit [RLU], glimepiride 1471±265 RLU, p<0.05).ConclusionLinagliptin is superior to glimepiride in regard to the preservation of endothelium-dependent relaxation in the presence of hyperglycemia and the improvement in endothelial function in response to linagliptin treatment is associated with greater antioxidant activity compared to glimepiride.

Ly6C+ Inflammatory Monocyte Differentiation Partially Mediates Hyperhomocysteinemia-Induced Vascular Dysfunction in Type 2 Diabetic db/db Mice.

Hyperhomocysteinemia (HHcy) is a potent risk factor for diabetic cardiovascular diseases. We have previously reported that hyperhomocysteinemia potentiates type 1 diabetes mellitus-induced inflammatory monocyte differentiation, vascular dysfunction, and atherosclerosis. However, the effects of hyperhomocysteinemia on vascular inflammation in type 2 diabetes mellitus (T2DM) and the underlying mechanism are unknown. Approach and Results: Here, we demonstrate that hyperhomocysteinemia was induced by a high methionine diet in control mice (homocysteine 129 µmol/L), which was further worsened in T2DM db/db mice (homocysteine 180 µmol/L) with aggravated insulin intolerance. Hyperhomocysteinemia potentiated T2DM-induced mononuclear cell, monocyte, inflammatory monocyte (CD11b+Ly6C+), and M1 macrophage differentiation in periphery and aorta, which were rescued by folic acid-based homocysteine-lowering therapy. Moreover, hyperhomocysteinemia exacerbated T2DM-impaired endothelial-dependent aortic relaxation to acetylcholine. Finally, transfusion of bone marrow cells depleted for Ly6C by Ly6c shRNA transduction improved insulin intolerance and endothelial-dependent aortic relaxation in hyperhomocysteinemia+T2DM mice. Hyperhomocysteinemia potentiated systemic and vessel wall inflammation and vascular dysfunction partially via inflammatory monocyte subset induction in T2DM. Inflammatory monocyte may be a novel therapeutic target for insulin resistance, inflammation, and cardiovascular complications in hyperhomocysteinemia+T2DM.

Betulinic acid attenuates lipopolysaccharide-induced vascular hyporeactivity in the rat aorta by modulating Nrf2 antioxidative function

Betulinic acid (BA), a pentacyclic triterpenoid, has been reported to inhibit cardiovascular dysfunction under sepsis-induced oxidative stress. Nuclear factor erythroid-2 related factor-2 (Nrf2) is regarded as a key transcription factor regulating expression of endogenous antioxidative genes. To explore the preventive effects of BA against vascular hyporeactivity and the related antioxidative mechanism in sepsis, contraction and relaxation in aortas isolated from lipopolysaccharide (LPS)-challenged rats were performed. Male Sprague-Dawley rats were pretreated with brusatol (Bru, 0.4mg/kg/2days, i.p.), an inhibitor of Nrf2, and BA (10, 25, 50mg/kg/day, i.g.) for 3days and injected with LPS (10mg/kg, i.p.) at the 4th day. Rats were anesthetized and killed by cervical dislocation after they were treated with LPS for 4h. Thoracic aortas were immediately dissected out to determine contraction and relaxation using the organ bath system. Pro-inflammatory factors interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) and oxidative stress were measured in aortic tissues and plasma. mRNA expression of Nrf2-regulated antioxidative enzymes, including superoxide dismutase (SOD), glutathione peroxidase (GPx), and heme oxygenase-1 (HO-1), in rat aortas was determined. Increases of IL-1β, TNF-α, nitric oxide, and malondialdehyde and the decrease of glutathione induced by LPS were significantly attenuated by pretreatment with different doses of BA in plasma and aortas (p < 0.05 versus LPS), all of which were blocked by Bru (p < 0.01). Inhibition of phenylephrine (PE)- and KCl-induced contractions and acetylcholine (ACh)-induced vasodilatation in aortas from LPS-challenged rats was dose-dependently reduced by BA (p < 0.05; percentage improvements by BA in PE-induced contraction were 55.38%, 96.41%, and 104.33%; those in KCl-induced contraction were 15.11%, 23.96%, and 22.96%; and those in ACh-induced vasodilatation were 16.08%, 42.99%, and 47.97%), all of which were reversed by Bru (p < 0.01). Improvements of SOD, GPx, and HO-1 mRNA expression conferred by BA in LPS-challenged rat aortas were inhibited by Bru (p < 0.01; 145.45% versus 17.42%, 160.69% versus 22.76%, and 166.88% versus 23.57%). These findings suggest that BA attenuates impairments of aortic contraction and relaxation in LPS-challenged rats by activating Nrf2-regulated antioxidative pathways.

Exercise training prevents the perivascular adipose tissue-induced aortic dysfunction with metabolic syndrome

The aim of the study was to determine the effects of exercise training on improving the thoracic perivascular adipose tissue (tPVAT) phenotype (inflammation, oxidative stress, and proteasome function) in metabolic syndrome and its subsequent actions on aortic function. MethodsLean and obese (model of metabolic syndrome) Zucker rats (n=8/group) underwent 8-weeks of control conditions or treadmill exercise (70% of max speed, 1 h/day, 5 days/week). At the end of the intervention, the tPVAT was removed and conditioned media was made. The cleaned aorta was attached to a force transducer to assess endothelium-dependent and independent dilation in the presence or absence of tPVAT-conditioned media. tPVAT gene expression, inflammatory /oxidative phenotype, and proteasome function were assessed. ResultsThe main findings were that Ex induced: (1) a beige-like, anti-inflammatory tPVAT phenotype; (2) a greater abundance of •NO in tPVAT; (3) a reduction in tPVAT oxidant production; and (4) an improved tPVAT proteasome function. Regarding aortic function, endothelium-dependent dilation was greater in exercised lean and obese groups vs. controls (p < 0.05). Lean control tPVAT improved aortic relaxation, whereas obese control tPVAT decreased aortic relaxation. In contrast, the obese Ex-tPVAT increased aortic dilation, whereas the lean Ex-tPVAT did not affect aortic dilation. ConclusionOverall, exercise had the most dramatic impact on the obese tPVAT reflecting a change towards an environment with less oxidant load, less inflammation and improved proteasome function. Such beneficial changes to the tPVAT micro-environment with exercise likely played a significant role in mediating the improvement in aortic function in metabolic syndrome following 8 weeks of exercise.

Epithelial sodium channels in endothelial cells mediate diet-induced endothelium stiffness and impaired vascular relaxation in obese female mice

ObjectiveMineralocorticoid receptor activation of the epithelial sodium channel in endothelial cells (ECs) (EnNaC) is accompanied by aldosterone induced endothelial stiffening and impaired nitric oxide (NO)-mediated arterial relaxation. Recent data support enhanced activity of the alpha subunit of EnNaC (αEnNaC) mediates this aldosterone induced endothelial stiffening and associated endothelial NO synthase (eNOS) activation. There is mounting evidence that diet induced obesity diminishes expression and activation of AMP-activated protein kinase α (AMPKα), sirtuin 1 (Sirt1), which would be expected to lead to impaired downstream eNOS activation. Thereby, we posited that enhanced EnNaC activation contributes to diet induced obesity related increases in stiffness of the endothelium and diminished NO mediated vascular relaxation by increasing oxidative stress and related inhibition of AMPKα, Sirt1, and associated eNOS inactivation. Materials/MethodsSixteen to twenty week-old αEnNaC knockout (αEnNaC−/−) and wild type littermate (EnNaC+/+) female mice were fed a mouse chow or an obesogenic western diet (WD) containing excess fat (46%) and fructose (17.5%) for 16 weeks. Sodium currents of ECs, endothelial stiffness and NO mediated aortic relaxation were examined along with indices of aortic oxidative stress, vascular remodeling and fibrosis. ResultsEnhanced EnNaC activation-mediated WD-induced increases in sodium currents in isolated lung ECs, increased endothelial stiffness and impaired aortic endothelium-dependent relaxation to acetylcholine (10−9–10−4 mol/L). These abnormalities occurred in conjunction with WD-mediated aortic tissue oxidative stress, inflammation, and decreased activation of AMPKα, Sirt1, and downstream eNOS were substantially mitigated in αEnNaC−/− mice. Importantly, αEnNaC−/− prevented WD induced increases in endothelial stiffness and related impairment of endothelium-dependent relaxation as well as aortic fibrosis and remodeling. However, EnNaC signaling was not involved in diet-induced abnormal expression of adipokines and CYP11b2 in abdominal aortic perivascular adipose tissue. ConclusionThese data suggest that endothelial specific EnNaC activation mediates WD-induced endothelial stiffness, impaired eNOS activation, aortic fibrosis and remodeling through increased aortic oxidative stress and increased inflammation related to a reduction of AMPKα and Sirt 1 mediated eNOS phosphorylation/activation and NO production.

Endothelium-dependent and independent effect of Guibourtia tessmannii (Caesalpiniaceae) on vascular contractility of rat

The stem barks of Guibourtia tessmannii (Caesalpiniaceae) are used in traditional Gabonese medicine as antihypertensive remedies. In the present study, we investigated vasorelaxant properties effect of aqueous extract from G. tessmannii and fuller understanding these mechanisms of action in vitro. The activity of Guibourtia tessmannii was evaluated on isolated aorta rings of rat constricted with KCl (80 mM) and norepinephrine (10-4 M). Cumulative concentrations (1 mg/mL - 100 mg/ml) of G. tessmannii provoked a dose-dependent relaxation of the thoracic aorta precontracted by norepinephrine or KCl (95.69 ± 0.6% and 91.34 ± 4.90%, respectively). The vasorelaxant effect induced by G. tessmannii on the aorta precontracted by KCl was significant decreased in presence of Nω-nitro-L-arginine methyl ester (11.30 ± 4.3 %, p&lt;0.05), tétraéthylammonium (52.2 ± 9.20 %, p&lt;0.01). Indomethacin and atropine modify the vasorelaxant effect of plant extract (57.13± 6.9 %, p&lt;0.01 and 58.83± 5.9 %, p&lt;0.01, respectively. These results suggest that the vasorelaxant action of G.tessmannii was mediated via the muscarinic receptors. The direct effect of G.tessmannii to be mediated by alpha-adrenergic receptors and potassium channels.

ERK-containing microparticles from a diabetic mouse induce endothelial dysfunction.

Endothelial dysfunction is a hallmark of diabetic vascular complications. Microparticles (MPs) are small vesicles shed from the surface of blood and vascular cells that act as stimuli and during apoptosis. Circulating MPs of diabetic rats have been shown to induce endothelial dysfunction. However, the underlying mechanisms require further study. In this study, we investigated how MPs from diabetic mice affect endothelial function. MPs were collected from streptozotocin-induced diabetic mice and Institute of Cancer Research (ICR) mice as controls. The levels of MPs were assessed and characterized by flow cytometry, enzyme-linked immunosorbent assay and dot blotting. Normal mice aortas were incubated with MPs and expressions of enzymes and vascular relaxation were analyzed. We found that (1) circulating MPs level increased in diabetic mice; (2) MPs impaired endothelial-dependent relaxation in mice aorta, but diabetic mice-derived MPs (diabetes mellitus (DM) MPs) were easier to attach to the endothelial cells than were control MPs; (3) DM MPs had more extracellular signal-regulated kinase (ERK)1/2 than did control mice-derived MPs, and they induced ERK1/2 activation in mice aortas; (4) DM MPs decreased endothelial nitric oxide synthase (eNOS) in mice aortas, and eNOS was emitted from endothelial cells to blood in the shape of endothelial MPs. DM MPs significantly altered endothelial function by activation of ERK1/2, which might provide a therapeutic target for diabetic vascular complications.

Adverse organogenesis and predisposed long-term metabolic syndrome from prenatal exposure to fine particulate matter.

Exposure to fine particulate matter (PM) during pregnancy is associated with high risks of birth defects/fatality and adverse long-term postnatal health. However, limited mechanistic data are available to assess the detailed impacts of prenatal PM exposure. Here we evaluate fine PM exposure during pregnancy on prenatal/postnatal organogenesis in offspring and in predisposing metabolic syndrome for adult life. Between days 0 and 18 of gestation, two groups of adult female rats (n = 10 for each) were placed in a dual-exposure chamber device, one with clean ambient air (∼3 µg·m-3) and the other with ambient air in the presence of 100 to 200 µg·m-3 of ultrafine aerosols of ammonium sulfate. At birth (postnatal day 0, PND0), four males and four females were selected randomly from each litter to be nursed by dams, whereas tissues were collected from the remaining pups. At PND21, tissues were collected from two males and two females, whereas the remaining pups were fed either a high- or low-fat diet until PND105, when tissues were obtained for biochemical and physiological analyses. Maternal exposure to fine PM increased stillbirths; reduced gestation length and birth weight; increased concentrations of glucose and free fatty acids in plasma; enhanced lipid accumulation in the liver; and decreased endothelium-dependent relaxation of aorta. This lead to altered organogenesis and predisposed progeny to long-term metabolic defects in an age-, organ-, and sex-specific manner. Our results highlight the necessity to develop therapeutic strategies to remedy adverse health effects of maternal PM exposure on conceptus/postnatal growth and development.

Voluntary wheel running activates Akt/AMPK/eNOS signaling cascades without improving profound endothelial dysfunction in mice deficient in α-galactosidase A

Fabry disease is caused by loss of activity of the lysosomal hydrolase α-galactosidase A (GLA). Premature life-threatening complications in Fabry patients arise from cardiovascular disease, including stroke and myocardial infarction. Exercise training has been shown to improve endothelial dysfunction in various settings including coronary artery disease. However, the effects of exercise training on endothelial dysfunction in Fabry disease have not been investigated. Gla knockout mice were single-housed in a cage equipped with a voluntary wheel (EX) or no wheel (SED) for 12 weeks. Exercised mice ran 10 km/day on average during the voluntary running intervention (VR) period. Despite significantly higher food intake in EX than SED, body weights of EX and SED remained stable during the VR period. After the completion of VR, citrate synthase activity in gastrocnemius muscle was significantly higher in EX than SED. VR resulted in greater phosphorylation of Akt (S473) and AMPK (T172) in the aorta of EX compared to SED measured by western blot. Furthermore, VR significantly enhanced eNOS protein expression and phosphorylation at S1177 by 20% and 50% in the aorta of EX when compared with SED. Similarly, plasma nitrate and nitrite levels were 77% higher in EX than SED. In contrast, measures of anti- and pro-oxidative enzymes (superoxide dismutase and p67phox subunit of NADPH oxidase) and overall oxidative stress (plasma oxidized glutathione) were not different between groups. Although the aortic endothelial relaxation to acetylcholine was slightly increased in EX, it did not reach statistical significance. This study provides the first evidence that VR improves Akt/AMPK/eNOS signaling cascades, but not endothelial function in the aorta of aged Gla deficient mice.

Moringa oleifera Seeds Improve Aging-Related Endothelial Dysfunction in Wistar Rats

Vascular aging is characterized by functional and structural changes of the vessel wall, including endothelial dysfunction, with decreased endothelial NO· bioavailability and elevated vasoconstrictor and inflammatory mediator production, vascular rigidity, and tone impairment. Moringa oleifera (MOI) is a little tree, and different parts of which are used in traditional medicine in tropical Africa, America, and Asia for therapeutic applications in several disorders including cardiovascular disease. The present study is aimed at assessing the effect of MOI on aging-associated alteration of the endothelial function in Wistar rats. Middle-aged Wistar rats (46-week-old males) have been fed with food containing or not 750 mg/kg/day of MOI seed powder for 4 weeks. A group of young Wistar rats (16-week-old) was used as control. Measurement of isometric contraction, western blot analysis, and immunostaining has then been performed in the aortas and mesenteric arteries to assess the endothelium function. MOI treatment improved carbachol-induced relaxation in both aortas and mesenteric arteries of middle-aged rats. In the aortas, this was associated with an increased Akt signalling and endothelial NO synthase activation and a downregulation of arginase-1. In the mesenteric arteries, the improvement of the endothelial-dependent relaxation was related to an EDHF-dependent mechanism. These results suggest a vascular protective effect of MOI seeds against the vascular dysfunction that develops during aging through different mechanisms in conductance and resistance arteries.

Putative Nox2 inhibitors worsen homocysteine-induced impaired acetylcholine-mediated relaxation

Background and aimIncreased homocysteine (Hcy) is associated with coronary artery disease (CAD). Hcy increases reactive oxygen species (ROS) via NADPH oxidases (Nox), reducing acetylcholine-mediated vasorelaxation. We aimed to determine if putative Nox2 inhibitors prevent Hcy-impaired acetylcholine-mediated vasorelaxation. Methods and resultsNew Zealand White rabbit and wild-type (C57BL/6) and Nox2−/− (NOX) mice aortic rings were mounted in organ baths. Rabbit rings were incubated with either apocynin (10 μM), gp91ds-tat (GP, 1 μM) or PhoxI2 (1 μM) and mice rings GP (1 μM) only. Some rabbit rings were incubated with 3 mM Hcy, before pre-contraction, followed by dose–response relaxation to acetylcholine (ACh; 0.01μM-10μM). In rabbit rings treated with Hcy and GP, O2‾ donor pyrogallol (1 μM) or Akt activator SC79 (1 μM) was added 5 min before ACh. Mice rings were used to compare Nox2 deletion to normal acetylcholine-mediated relaxation.In rabbits, Hcy reduced acetylcholine-mediated relaxation vs. control (p < 0.0001). Treatment + Hcy reduced relaxation compared with treatment alone (p < 0.0001). Pyrogallol and SC79 reversed the response of GP + Hcy (p = 0.0001). In mice, Nox2 deletion reduced acetylcholine-mediated vasorelaxation. Rabbit tissue analysis revealed that Hcy reduced eNOS phosphorylation at Thr495 and increased eNOS phosphorylation at Ser1177; no further alteration at Thr495 was observed with GP. In contrast, GP prevented increased phosphorylation at Ser1177. ConclusionsApocynin, GP and PhoxI2 worsens acetylcholine-mediated vascular relaxation in rabbit aorta, which is supported by results from mouse Nox2 deletion data. These inhibitors worsen Hcy-induced vascular dysfunction, suggesting that current putative Nox2 inhibitors might not be useful in treating HHcy.

The cardiovascular effects of Aspalathus linearis supplementation in male Wistar rats receiving fixed-dose combination first-line antiretroviral therapy.

HIV-infected populations receiving antiretroviral therapy (ART) have an increased risk of cardiovascular disease. The beneficial cardiovascular effects of rooibos are well described; however, it is unknown whether rooibos ameliorates harmful ART-induced cardiovascular side effects. We investigated the cardiometabolic effects of rooibos co-treatment in rats receiving ART (efavirenz, emtricitabine, tenofovir) for nine weeks. Rooibos treatment reduced total cholesterol levels; however, triglyceride, phospholipid and thiobarbituric acidreactive substance levels were unaffected by ART, rooibos or combination treatment. In isolated hearts exposed to ischaemia-reperfusion injury, ART resulted in increased infarct sizes compared to controls, which was not observed when co-treated with rooibos. Vascular studies showed reduced aortic relaxation with ART, and improved relaxation when co-treated with rooibos. In conclusion, we show that rooibos treatment reduced total cholesterol levels in control rats, and that rooibos co-treatment ameliorated the harmful ART-induced cardiovascular effects. These findings are novel and warrant further studies into underlying mechanisms and clinical relevance.

Prenatal Sildenafil Therapy Improves Cardiovascular Function in Fetal Growth Restricted Offspring of Dahl Salt-Sensitive Rats.

Fetal growth restriction (FGR) is associated with increased risk for cardiovascular and renal disorders in later life. Prenatal sildenafil improves birth weight in FGR animal models. Whether sildenafil treatment protects against long-term cardiovascular and renal disease in these offspring is unknown. The aim of this study is to test the hypothesis that prenatal sildenafil ameliorates cardiovascular and renal function in FGR offspring of Dahl salt-sensitive rats. Sildenafil citrate (60 mg/kg per day) or control gel diet (containing 0.3% salt) was administered from gestational day ten until birth. In male and female offspring, the mean arterial pressure was measured by telemetry in 1 subset from week 5 until week twenty. Echocardiographic parameters, glomerular filtration rate, and fractional electrolyte excretion were determined in another subset at week 9. Aortic and mesenteric artery rings were prepared to assess endothelial-dependent (acetylcholine) and -independent (sodium nitroprusside) vasorelaxation (week 10). The rise in mean arterial pressure per week was attenuated in treated versus untreated male offspring. Mesenteric arteries showed an increased endothelium-dependent relaxation and improved endothelium-independent relaxation in treated versus control male offspring. No differences in aortic relaxation, echocardiographic parameters or renal function were observed between groups. Prenatal sildenafil treatment subtly improves cardiovascular but not renal function in the offspring of this FGR rat model. Translationally, in utero treatment could be beneficial for cardiovascular programming in a sex-specific manner; however, caution is warranted since recent human trials have been halted because of potentially deleterious neonatal side effects when treating pregnancies complicated with severe FGR with sildenafil.

Mulberry Extract Attenuates Endothelial Dysfunction through the Regulation of Uncoupling Endothelial Nitric Oxide Synthase in High Fat Diet Rats.

Dyslipidemia is associated with endothelial dysfunction, which is linked to nitric oxide (NO) biology. The coupling of endothelial NO synthase with cofactors is a major step for NO release. This study is aimed to investigate the vascular pharmacology effects of mulberry in rat thoracic aorta and human vascular endothelial cells. In vitro, we investigated the protective effects of the mulberry extract and its main component cyanidin-3-rutinoside (C-3-R), against oxidized low-density lipoprotein (ox-LDL)-induced endothelial nitric oxide synthase (eNOS) uncoupling. Whereas ox-LDL significantly decreased NO levels in endothelial cells, mulberry extract, and C-3-R significantly recovered NO levels and phospho-eNOS Thr495 and Ser1177 expression. In vivo, mulberry was administered to 60% of high-fat diet (w/w)-fed Sprague Dawley (SD) rats for six weeks, in which endothelium-dependent relaxations were significantly improved in organ bath studies and isometric tension recordings. Consistently, aortic expressions of phospho-eNOS and nitrotyrosine were increased. Mulberry also raised serum NO levels, increased phosphorylation of eNOS, and reduced nitrotyrosine and intracellular reactive oxygen species (ROS) in aortas, showing that mulberry preserves endothelium-dependent relaxation in aortas from high-fat diet rats. We suggest that this effect is mediated through enhanced NO bioavailability, in which the regulation of ROS and its reduced eNOS uncoupling are involved.

L-cysteine/cystathionine-β-synthase-induced relaxation in mouse aorta involves a L-serine/sphingosine-1-phosphate/NO pathway.

Among the three enzymes involved in the transsulfuration pathway, only cystathionine β-synthase (CBS) converts L-cysteine into L-serine and H2 S. L-serine is also involved in the de novo sphingolipid biosynthesis through a condensation with palmitoyl-CoA by the action of serine palmitoyltransferase (SPT). Here, we have investigated if L-serine contributes to the vasorelaxant effect. The presence of CBS in mouse vascular endothelium was assessed by immunohistochemistry and immunofluorescence. The relaxant activity of L-serine (0.1-300μM) and L-cysteine (0.1-300μM) was estimated on mouse aorta rings, with or without endothelium. A pharmacological modulation study evaluated NO and sphingosine-1-phosphate (S1P) involvement. Levels of NO and S1P were also measured following incubation of aorta tissue with either L-serine (1, 10, and 100μM) or L-cysteine (10, 100μM, and 1mM). L-serine relaxed aorta rings in an endothelium-dependent manner. The vascular effect was reduced by L-NG-nitro-arginine methyl ester and wortmaninn. A similar pattern was obtained with L-cysteine. The S1P1 receptor antagonist (W146) or the SPT inhibitor (myriocin) reduced either L-serine or L-cysteine relaxant effect. L-serine or L-cysteine incubation increased NO and S1P levels in mouse aorta. L-serine, a by-product formed within the transsulfuration pathway starting from L-cysteine via CBS, contributes to the vasodilator action of L-cysteine. The L-serine effect involves both NO and S1P. This mechanism could be involved in the marked dysregulation of vascular tone in hyperhomocysteinemic patients (CBS deficiency) and may represent a feasible therapeutic target. This article is part of a themed section on Hydrogen Sulfide in Biology & Medicine. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v177.4/issuetoc.

Sulodexide promotes arterial relaxation via endothelium-dependent nitric oxide-mediated pathway

Sulodexide (SDX) is a highly purified glycosaminoglycan with antithrombotic and profibrinolytic properties and reported benefits in thrombotic and atherosclerotic vascular disorders. However, the effects of SDX on vascular function are unclear. We tested whether SDX affects vascular relaxation and examined the potential underlying mechanisms. Isolated segments of male rat abdominal aorta and mesenteric artery were suspended in a tissue bath, and the changes in arterial contraction/relaxation were measured. The α-adrenergic receptor agonist phenylephrine (Phe) (10−9–10−5 M) caused concentration-dependent aortic and mesenteric artery contraction that was reduced in tissues pretreated with SDX (1 mg/ml). In aortic and mesenteric arterial segments precontracted with submaximal concentration of Phe (3 × 10−7–6 × 10−7 M), SDX (0.001–1 mg/ml) caused concentration-dependent relaxation. To test the role of endothelium, SDX-induced relaxation was compared with that of acetylcholine (ACh), a known activator of endothelium-dependent relaxation. In Phe precontracted aorta, ACh relaxation was abolished and SDX relaxation was significantly inhibited by endothelium removal or the nitric oxide synthase (NOS) inhibitor Nω-nitro-l-arginine methyl ester (L-NAME), suggesting a role of NO. In mesenteric artery, ACh relaxation was abolished by endothelium removal, partially blocked by L-NAME, and completely blocked by a mixture of indomethacin, a cyclooxygenase inhibitor and blocker of the PGI2-cAMP pathway, and tetraethylammonium, a blocker of K+ channels and EDHF-dependent hyperpolarization pathway. In comparison, SDX relaxation of mesenteric artery was almost completely inhibited by endothelium removal or NOS inhibitor L-NAME. SDX enhanced vascular relaxation and increased nitrate/nitrite production in response to all ACh concentrations in the aorta, but only to low ACh concentrations (<10−7 M) in mesenteric artery. SDX did not affect aortic or mesenteric artery endothelium-independent relaxation to the NO donor sodium nitroprusside. Thus, SDX promotes arterial relaxation via a mechanism involving endothelium-dependent NO production; an effect that could enhance vasodilation and decrease vasoconstriction in vascular disorders.

Human genotyping and an experimental model reveal NPR-C as a possible contributor to morbidity in coarctation of the aorta.

Coarctation of the aorta (CoA) is a common congenital cardiovascular (CV) defect characterized by a stenosis of the descending thoracic aorta. Treatment exists, but many patients develop hypertension (HTN). Identifying the cause of HTN is challenging because of patient variability (e.g., age, follow-up duration, severity) and concurrent CV abnormalities. Our objective was to conduct RNA sequencing of aortic tissue from humans with CoA to identify a candidate gene for mechanistic studies of arterial dysfunction in a rabbit model of CoA devoid of the variability seen with humans. We present the first known evidence of natriuretic peptide receptor C (NPR-C; aka NPR3) downregulation in human aortic sections subjected to high blood pressure (BP) from CoA versus normal BP regions (validated to PCR). These changes in NPR-C, a gene associated with BP and proliferation, were replicated in the rabbit model of CoA. Artery segments from this model were used with human aortic endothelial cells to reveal the functional relevance of altered NPR-C activity. Results showed decreased intracellular calcium ([Ca2+]i) activity to C-type natriuretic peptide (CNP). Normal relaxation induced by CNP and atrial natriuretic peptide was impaired for aortic segments exposed to elevated BP from CoA. Inhibition of NPR-C (M372049) also impaired aortic relaxation and [Ca2+]i activity. Genotyping of NPR-C variants predicted to be damaging revealed that rs146301345 was enriched in our CoA patients, but sample size limited association with HTN. These results may ultimately be used to tailor treatment for CoA based on mechanical stimuli, genotyping, and/or changes in arterial function.

Endothelium-independent vasodilator effects of nobiletin in rat aorta

Nobiletin is a one of the polymethoxyflavones contained in the peel of citrus fruits, such as Citrus depressa. In this study, the effect of nobiletin-induced relaxation on phenylephrine (PE)-induced contraction of endothelium-denuded rat aorta was investigated. Nobiletin inhibited PE- or KCl-induced contractions in a concentration-dependent manner in endothelium-intact and -denuded aortas. However, this relaxation was stronger in PE-induced contractions than in KCl-induced contractions; moreover, the nobiletin-induced relaxation was significantly increased on PE-induced contraction in endothelium-intact aorta. ODQ significantly inhibited the nobiletin-induced relaxation in endothelium-denuded aorta; however, SQ22536 did not affect the relaxation. In addition, IBMX synergistically enhanced the nobiletin-induced relaxation. Nobiletin increased cGMP levels in aorta. Also, IBMX significantly increased cGMP content in aorta, and ODQ significantly reduced cGMP levels. Nobiletin-induced relaxation was significantly inhibited by the Ca2+-activated K+ (BK) channel inhibitor iberiotoxin (IbTX) and the ATP-sensitive K+ (KATP) channel inhibitor glybenclamide. Sodium nitroprusside-induced relaxation was suppressed by IbTX, but not by glybenclamide. These results suggest that nobiletin inhibits PE-induced contractions of endothelium-denuded rat aorta by increasing cGMP levels via GC activation. Moreover, the present findings indicate the possibility that nobiletin opened BK channels by a cGMP-related signal, but KATP channels were opened by a cGMP-nonrelated signal in rat aorta.