The non-obligatory role of endothelial cells in the relaxation of arterial smooth muscle by acetylcholine

The concept of Endothelium Derived Relaxing Factor (EDRF), put forward by Furchgott in the earlier 80s of the past century, implies that nitric oxide (NO) produced by NO synthase (NOS) in the endothelium in response to acetylcholine (ACh) passively diffuses to the underlying vascular smooth muscle cells (VSMC) thereby reducing vascular tension. It was thought that VSMC do not express NOS by themselves, but to the time of those studies immunohistochemical techniques were not what they are now. State-of-the-art immunohistochemistry permits nowadays to localize NOS both to the endothelium and to VSMC. However, the principal question remained unanswered, is the NO generation by VSMC physiologically relevant? We hypothesized that the destruction of the vascular wall anatomical integrity by rubbing the blood vessel intimal surface may increase vascular superoxides that, in turn, reduce NO bioactivity. To address this issue, we examined ACh-induced vasorelaxation in endothelium-deprived blood vessels under protection against oxidative stress and found that superoxide scavengers - tempol and N-acetyl-L-cysteine - restored vasodilatory responses to ACh in endothelium-deprived blood vessels without influencing the vascular wall tension in intact blood vessels. Herewith we provided the first evidence that VSMC can release NO in amounts sufficient to account for the vasorelaxatory response to ACh. In contrast to the commonly accepted concept of the obligatory role of endothelial cells in the relaxation of arterial smooth muscle, the local NO generation by VSMC can modulate vascular functions in an endothelium-independent manner

The Effect of Chronic NO Synthase Inhibition on the Vasoactive and Structural Properties of Thoracic Aorta, NO Synthase Activity, and Oxidative Stress Biomarkers in Young SHR.

Although the role of nitric oxide (NO) in essential hypertension is still unclear, the effects of long-term NO deficiency have not yet been investigated during the critical juvenile period in spontaneously hypertensive rats (SHR). We aimed to analyze the effects of chronic NO synthase (NOS) inhibition on systolic blood pressure (sBP), vasoactivity, morphological changes and superoxide level in the thoracic aorta (TA), NOS activity in different tissues, and general biomarkers of oxidative stress in plasma of young SHR. Four-week-old SHR were treated with NG-nitro-L-arginine methyl ester (L-NAME, 50 mg/kg/day, p.o.) for 4-5 weeks. L-NAME treatment induced a transient sBP increase only, and surprisingly, slightly inhibited endothelium-dependent relaxation of TA. Hereby, the inhibition of NOS activity varied from tissue to tissue, ranging from the lowest in the TA and the kidney to the highest in the brain stem. In spite of an increased sensitivity of adrenergic receptors, the maximal adrenergic contraction of TA was unchanged, which was associated with changes in elastin arrangement and an increase in wall thickness. The production of reactive oxygen species in the TA was increased; however, the level of selected biomarkers of oxidative stress did not change. Our findings proved that the TA of young SHR responded to chronic NO deficiency by the development of adaptive mechanisms on the functional (preserved NO-derived vasorelaxation, unincreased contraction) and molecular (preserved NOS activity) level

AVE0991, a nonpeptide angiotensin 1-7 receptor agonist, improves glucose metabolism in the skeletal muscle of obese zucker rats : possible involvement of prooxidant/antioxidant mechanisms

Angiotensin 1-7 (Ang 1-7) enhances insulin signaling and glucose transport activity in the skeletal muscle. The aim of our study was to evaluate the effect of AVE0991, a nonpeptide Mas receptor agonist, on the metabolic parameters, expression of RAS components and markers of oxidative stress, and insulin signaling in the skeletal morbidly obese rats. 33-week-old male obese Zucker rats were treated with vehicle and AVE0991 (0.5 mg/kg BW/day) via osmotic minipumps for two weeks. Gene expressions were determined by qPCR and/or Western blot analysis in musculus quadriceps. The enzymatic activities were detected flourometrically (aminopeptidase A) or by colorimetric assay kit (protein tyrosine phosphatase 1B). Administration of AVE0991 enhanced insulin signaling cascade in the skeletal muscle, reflected by improved whole-body glucose tolerance. It has been shown that reactive oxygen species (ROS) have insulin-mimetic action in muscle. The expression of renin receptor, transcription factor PLZF, and prooxidant genes was upregulated by AVE0991 accompanied by elevated expression of genes coding enzymes with antioxidant action. Our results show that AVE0991 administration activates genes involved in both ROS generation and clearance establishing a new prooxidant/antioxidant balance on a higher level, which might contribute to the improved insulin signaling pathway and glucose tolerance of obese Zucker rats

The reaction products of sulfide and S-nitrosoglutathione are potent vasorelaxants

The chemical interaction of sodium sulfide (Na2S) with the NO-donor S-nitrosoglutathione (GSNO) has been described to generate new reaction products, including polysulfides and nitrosopersulfide (SSNO-) via intermediacy of thionitrous acid (HSNO). The aim of the present work was to investigate the vascular effects of the longer-lived products of the Sulfide/GSNO interaction. Here we show that the products of this reaction relax precontracted isolated rings of rat thoracic aorta and mesenteric artery (but to a lesser degree rat uterus) with a >2-fold potency compared with the starting material, GSNO (50?nM), whereas Na2S and polysulfides have little effect at 1-5?µM. The onset of vasorelaxation of the reaction products was 7-10 times faster in aorta and mesenteric arteries compared with GSNO. Relaxation to GSNO (100-500?nM) was blocked by an inhibitor of soluble guanylyl cyclase, ODQ (0.1 and 10?µM), and by the NO scavenger cPTIO (100?µM), but less affected by prior acidification (pH 2-4), and unaffected by N-acetylcysteine (1?mM) or methemoglobin (20?µM heme). By contrast, relaxation to the Sulfide/GSNO reaction products (100-500?nM based on the starting material) was inhibited to a lesser extent by ODQ, only slightly decreased by cPTIO, more markedly inhibited by methemoglobin and N-acetylcysteine, and abolished by acidification before addition to the organ bath. The reaction mixture was found to generate NO as detected by EPR spectroscopy using N-(dithiocarboxy)-N-methyl-D-glucamine (MGD2)-Fe2+ as spin trap. In conclusion, the Sufide/GSNO reaction products are faster and more pronounced vasorelaxants than GSNO itself. We conclude that in addition to NO formation from SSNO-, reaction products other than polysulfides may give rise to nitroxyl (HNO) and be involved in the pronounced relaxation induced by the Sulfide/GSNO cross-talk

Arterial Hypertension and Plasma Glucose Modulate the Vasoactive Effects of Nitroso-Sulfide Coupled Signaling in Human Intrarenal Arteries

We have investigated the vasoactive effects of the coupled nitro-sulfide signaling pathway in lobar arteries (LAs) isolated from the nephrectomized kidneys of cancer patients: normotensive patients (NT) and patients with arterial hypertension (AH). LAs of patients with AH revealed endothelial dysfunction, which was associated with an increased response to the exogenous NO donor, nitrosoglutathione (GSNO). The interaction of GSNO with the H2S donor triggered a specific vasoactive response. Unlike in normotensive patients, in patients with AH, the starting and returning of the vasorelaxation induced by the end-products of the H2S-GSNO interaction (S/GSNO) was significantly faster, however, without the potentiation of the maximum. Moreover, increasing glycemia shortened the time required to reach 50% of the maximum vasorelaxant response induced by S/GSNO products so modulating their final effect. Moreover, we found out that, unlike K+ channel activation, cGMP pathway and HNO as probable mediator could be involved in mechanisms of S/GSNO action. For the first time, we demonstrated the expression of genes coding H2S-producing enzymes in perivascular adipose tissue and we showed the localization of these enzymes in LAs of normotensive patients and in patients with AH. Our study confirmed that the heterogeneity of specific nitroso-sulfide vasoactive signaling exists depending on the occurrence of hypertension associated with increased plasma glucose level. Endogenous H2S and the end-products of the H2S-GSNO interaction could represent prospective pharmacological targets to modulate the vasoactive properties of human intrarenal arteries

The effect of the long-term inhibition of hydrogen sulfide production on the reactivity of the cardiovascular system in Wistar rats.

In this study, we investigated the blood pressure responses of the peripheral bed in vivo after chronic hydrogen sulphide (H2S) inhibition combined with acute nitric oxide (NO) deficiency. We also evaluated the role of endogenously produced H2S in the vasoactive responses of large- and medium-sized arteries in vitro. Changes in integrated blood pressure responses were measured after chronic inhibition of cystathionine-γ-lyase, an enzyme involved in H2S synthesis, with DL-propargylglycine (PPG), and acute inhibition of NO-synthase with nonspecific L-NG-nitro arginine methyl ester (L-NAME), and vasoactive responses of the thoracic aorta (TA) and mesenteric artery (MA) were investigated after acute incubation with PPG. We confirmed that chronic H2S deficiency had no effect on blood pressure, heart trophycity, noradrenaline and H2S donor vasoactive responses but induced renal hypertrophy and decrease in acetylcholine-induced hypotensive and L-NAME-induced hypertensive responses. Acute H2S deficiency led to an increase in basal tone (MA) or active tone (TA), whereas endothelium-dependent vasorelaxation remained unaffected. Long-term administration of PPG revealed a role of endogenous H2S in the bioavailability of endothelial NO in peripheral arteries. When both H2S and NO were lacking, the activation of H2S-independent compensatory mechanisms plays an important role in maintaining the vasodilator responses of the cardiovascular system.The accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author

Fructose Intake Impairs the Synergistic Vasomotor Manifestation of Nitric Oxide and Hydrogen Sulfide in Rat Aorta

In this study, we evaluated the effect of eight weeks of administration of 10% fructose solution to adult Wistar Kyoto (WKY) rats on systolic blood pressure (SBP), plasma and biometric parameters, vasoactive properties of the thoracic aorta (TA), NO synthase (NOS) activity, and the expression of enzymes producing NO and H2S. Eight weeks of fructose administration did not affect SBP, glycaemia, or the plasma levels of total cholesterol or low-density and high-density lipoprotein; however, it significantly increased the plasma levels of γ-glutamyl transferase and alanine transaminase. Chronic fructose intake deteriorated endothelium-dependent vasorelaxation (EDVR) and increased the sensitivity of adrenergic receptors to noradrenaline. Acute NOS inhibition evoked a reduction in EDVR that was similar between groups; however, it increased adrenergic contraction more in fructose-fed rats. CSE inhibition decreased EDVR in WKY but not in fructose-fed rats. The application of a H2S scavenger evoked a reduction in the EDVR in WKY rats and normalized the sensitivity of adrenergic receptors in rats treated with fructose. Fructose intake did not change NOS activity but reduced the expression of eNOS and CBS in the TA and CSE and CBS in the left ventricle. Based on our results, we could assume that the impaired vascular function induced by increased fructose intake was probably not directly associated with a decreased production of NO, but rather with impairment of the NO–H2S interaction and its manifestation in vasoactive responses

The Vasoactive Effect of Perivascular Adipose Tissue and Hydrogen Sulfide in Thoracic Aortas of Normotensive and Spontaneously Hypertensive Rats

The objective of this study was to investigate the vasoregulatory role of perivascular adipose tissue (PVAT) and its mutual interaction with endogenous and exogenous H2S in the thoracic aorta (TA) of adult normotensive Wistar rats and spontaneously hypertensive rats (SHRs). In SHRs, hypertension was associated with cardiac hypertrophy and increased contractility. Regardless of the strain, PVAT revealed an anticontractile effect; however, PVAT worsened endothelial-dependent vasorelaxation. Since H2S produced by both the vascular wall and PVAT had a pro-contractile effect in SHRs, H2S decreased the sensitivity of adrenergic receptors to noradrenaline in Wistar rats. While H2S had no contribution to endothelium-dependent relaxation in Wistar rats, in SHRs, H2S produced by the vascular wall had a pro-relaxant effect. We observed a larger vasorelaxation induced by exogenous H2S donor in SHRs than in Wistar rats. Additionally, in the presence of PVAT, this effect was potentiated. We demonstrated that PVAT of the TA aggravated endothelial function in SHRs. However, H2S produced by the TA vascular wall had a pro-relaxation effect, and PVAT revealed anti-contractile activity mediated by the release of an unknown factor and potentiated the vasorelaxation induced by exogenous H2S. All these actions could represent a form of compensatory mechanism to balance impaired vascular tone regulation