Chronic endothelial dopamine receptor stimulation improves endothelial function and hemodynamics in autosomal dominant polycystic kidney disease.

Altered polycystin-mediated endothelial flow mechanosensitivity contributes to the development of hypertension and cardiovascular complications in patients with autosomal dominant polycystic kidney disease (ADPKD). Stimulation of endothelial type 5 dopamine receptors (DR5) can acutely compensate for the endothelial consequences of polycystin deficiency, but the chronic impact of this approach must be evaluated in ADPKD. Nineteen patients with ADPKD on standard of care therapy were randomized to receive a 2-month treatment with the DR agonist rotigotine using transdermal patches, nine at 2 mg/24hours and ten at 4 mg/24hours or while ten were untreated. Rotigotine at the dose of 4 mg/24hours significantly increased nitric oxide release (nitrite levels from 10±30 to 46±34 nmol/L) and radial artery endothelium-dependent flow-mediated dilatation (from 16.4±6.3 to 22.5±7.3%) in response to hand skin heating. Systemic hemodynamics were not significantly modified but aplanation tonometry showed that rotigotine at 4 mg/24hours reduced aortic augmentation index and pulse pressure without affecting carotid-to femoral pulse wave velocity. Plasma creatinine and urea, urinary cyclic AMP, which contributes to cyst growth in ADPKD and copeptin, a surrogate marker of vasopressin, were not affected by rotigotine. In mice with a specific deletion of polycystin-1 in endothelial cells, chronic infusion of the peripheral DR5 agonist fenoldopam also improved mesenteric artery flow-mediated dilatation and reduced blood pressure. Thus, our study demonstrates that in patients with ADPKD, chronic administration of rotigotine improves conduit artery endothelial function through the restoration of flow-induced nitric oxide release as well as hemodynamics suggesting that endothelial DR5 activation may represent a promising pharmacological approach to prevent cardiovascular complications of ADPKD.

Altered bioavailability of epoxyeicosatrienoic acids is associated with conduit artery endothelial dysfunction in type 2 diabetic patients.

BACKGROUND: This pathophysiological study addressed the hypothesis that soluble epoxide hydrolase (sEH), which metabolizes the vasodilator and anti-inflammatory epoxyeicosatrienoic acids (EETs) to dihydroxyeicosatrienoic acids (DHETs), contributes to conduit artery endothelial dysfunction in type 2 diabetes. METHODS AND RESULTS: Radial artery endothelium-dependent flow-mediated dilatation in response to hand skin heating was reduced in essential hypertensive patients (n = 9) and type 2 diabetic subjects with (n = 19) or without hypertension (n = 10) compared to healthy subjects (n = 36), taking into consideration cardiovascular risk factors, flow stimulus and endothelium-independent dilatation to glyceryl trinitrate. Diabetic patients but not non-diabetic hypertensive subjects displayed elevated whole blood reactive oxygen species levels and loss of NO release during heating, assessed by measuring local plasma nitrite variation. Moreover, plasma levels of EET regioisomers increased during heating in healthy subjects, did not change in hypertensive patients and decreased in diabetic patients. Correlation analysis showed in the overall population that the less NO and EETs bioavailability increases during heating, the more flow-mediated dilatation is reduced. The expression and activity of sEH, measured in isolated peripheral blood mononuclear cells, was elevated in diabetic but not hypertensive patients, leading to increased EETs conversion to DHETs. Finally, hyperglycemic and hyperinsulinemic euglycemic clamps induced a decrease in flow-mediated dilatation in healthy subjects and this was associated with an altered EETs release during heating. CONCLUSIONS: These results demonstrate that an increased EETs degradation by sEH and altered NO bioavailability are associated with conduit artery endothelial dysfunction in type 2 diabetic patients independently from their hypertensive status. The hyperinsulinemic and hyperglycemic state in these patients may contribute to these alterations. Trial registration NCT02311075. Registered December 8, 2014.

A sensitive LC-MS/MS method for the quantification of regioisomers of epoxyeicosatrienoic and dihydroxyeicosatrienoic acids in human plasma during endothelial stimulation.

Epoxyeicosatrienoic acids (EETs) are vasodilating lipid mediators metabolized into dihydroxyeicosatrienoic acids (DHETs) by soluble epoxide hydrolase. We aimed to develop a LC-MS/MS method to quantify EETs and DHETs in human plasma and monitored their levels during vascular endothelial stimulation. Plasma samples, collected from 14 healthy and five hypertensive subjects at baseline and during radial artery endothelium-dependent flow-mediated dilatation, were spiked with internal standards. Lipids were then extracted by a modified Bligh and Dyer method and saponified to release bound EETs and DHETs. Samples were purified by a second liquid-liquid extraction and analyzed by LC-MS/MS. The assay allowed identification of (±)8(9)-epoxy-5Z,11Z,14Z-eicosatrienoic acid (8,9-EET); (±)11(12)-epoxy-5Z,8Z,14Z-eicosatrienoic acid (11,12-EET); (±)14(15)-epoxy-5Z,8Z,11Z-eicosatrienoic acid (14,15-EET); (±)8,9-dihydroxy-5Z,11Z,14Z-eicosatrienoic acid (8,9-DHET); (±)11,12-dihydroxy-5Z,8Z,14Z-eicosatrienoic acid (11,12-DHET); and (±)14,15-dihydroxy-5Z,8Z,11Z-eicosatrienoic acid (14,15-DHET). (±)5(6)-epoxy-5Z,11Z,14Z-eicosatrienoic acid (5,6-EET) was virtually undetectable due to its chemical instability. The limits of quantification were 0.25 ng/mL for DHETs and 0.5 ng/mL for EETs. Intra- and inter-assay variations ranged from 1.6 to 13.2%. Heating induced a similar increase in 8,9-EET, 11,12-EET, and 14,15-EET levels and in corresponding DHET levels in healthy but not in hypertensive subjects. We validated a sensitive LC-MS/MS method for measuring simultaneously plasma EET and DHET regioisomers in human plasma and showed its interest for assessing endothelial function.

Polycystin deficiency induces dopamine-reversible alterations in flow-mediated dilatation and vascular nitric oxide release in humans.

Autosomal dominant polycystic kidney disease (ADPKD) is a renal hereditary disorder associated with increased cardiovascular mortality, due to mutations in polycystin-1 and polycystin-2 genes. Endothelial polycystin-deficient cells have an altered mechanosensitivity to fluid shear stress and subsequent deficit in calcium-induced nitric oxide release, prevented by dopamine receptor stimulation. However, the impact of polycystin deficiency on endothelial function in ADPKD patients is still largely unknown. Here we assessed endothelium-dependent flow-mediated dilatation in 21 normotensive ADPKD patients and 21 healthy control subjects, during sustained (hand skin heating) and transient (postischemic hyperemia) flow stimulation. Flow-mediated dilatation was less marked in ADPKD patients than in controls during heating, but it was similar during postischemic hyperemia. There was no difference in endothelium-independent dilatation in response to glyceryl trinitrate. Local plasma nitrite, an indicator of nitric oxide availability, increased during heating in controls but not in patients. Brachial infusion of dopamine in a subset of ADPKD patients stimulated plasma nitrite increase during heating and improved flow-mediated dilatation. Thus, ADPKD patients display a loss of nitric oxide release and an associated reduction in endothelium-dependent dilatation of conduit arteries during sustained blood flow increase. The correction of these anomalies by dopamine suggests future therapeutic strategies that could reduce the occurrence of cardiovascular events in ADPKD.

High-efficiency on-line haemodiafiltration improves conduit artery endothelial function compared with high-flux haemodialysis in end-stage renal disease patients.

BACKGROUND: Middle molecular weight uraemic toxins are considered to play an important role in vascular dysfunction and cardiovascular outcomes in end-stage renal disease (ESRD) patients. Recent dialysis techniques based on convection, specifically high-efficiency on-line haemodiafiltration (HDF), enhance the removal of middle molecular weight toxins and reduce all-cause mortality in haemodialysis (HD) patients. However, the mechanisms of these improved outcomes remain to be established. METHODS: This prospective study randomly assigned 42 ESRD patients to switch from high-flux HD to high-efficiency on-line HDF (n=22) or to continue HD (n=20). Brachial artery endothelium-dependent flow-mediated dilatation, central pulse pressure, carotid artery intima-media thickness (IMT), internal diastolic diameter and distensibility and circulating markers of uraemia, inflammation and oxidative stress were blindly assessed before and after a 4-month follow-up. RESULTS: Brachial flow-mediated dilatation and carotid artery distensibility increased significantly in the HDF group compared with HD, while carotid IMT and diameter remained similar. HDF decreased predialysis levels of the uraemic toxins ß2-microglobulin, phosphate and blood TNFα mRNA expression. Oxidative stress markers were not different between the HD and HDF groups. Blood mRNA expression of protein kinase C ß2, an endothelial NO-synthase (eNOS) inhibitor, decreased significantly with HDF. CONCLUSIONS: High-efficiency on-line HDF prevents the endothelial dysfunction and stiffening of the conduit arteries in ESRD patients compared with high-flux HD. HDF decreases uraemic toxins, vascular inflammation, and is associated with subsequent improvement in eNOS functionality. These results suggest that reduced endothelial dysfunction may be an intermediate mechanism explaining the beneficial outcomes associated with HDF.

Epoxyeicosatrienoic acids contribute with altered nitric oxide and endothelin-1 pathways to conduit artery endothelial dysfunction in essential hypertension.

BACKGROUND: We sought to clarify, using functional and biological approaches, the role of epoxyeicosatrienoic acids, nitric oxide (NO)/reactive oxygen species balance, and endothelin-1 in conduit artery endothelial dysfunction during essential hypertension. METHODS AND RESULTS: Radial artery diameter and mean wall shear stress were determined in 28 untreated patients with essential hypertension and 30 normotensive control subjects during endothelium-dependent flow-mediated dilatation induced by hand skin heating. The role of epoxyeicosatrienoic acids and NO was assessed with the brachial infusion of inhibitors of cytochrome P450 epoxygenases (fluconazole) and NO synthase (N(G)-monomethyl-l-arginine [L-NMMA]). Compared with controls, hypertensive patients exhibited a decreased flow-mediated dilatation in response to postischemic hyperemia as well as to heating, as shown by the lesser slope of their diameter-shear stress relationship. In controls, heating-induced flow-mediated dilatation was reduced by fluconazole, L-NMMA, and, to a larger extent, by L-NMMA+fluconazole. In patients, flow-mediated dilatation was not affected by fluconazole and was reduced by L-NMMA and L-NMMA+fluconazole to a lesser extent than in controls. Furthermore, local plasma epoxyeicosatrienoic acids increased during heating in controls (an effect diminished by fluconazole) but not in patients. Plasma nitrite, an indicator of NO availability, increased during heating in controls (an effect abolished by L-NMMA) and, to a lesser extent, in patients, whereas, inversely, reactive oxygen species increased more in patients (an effect diminished by L-NMMA). Plasma endothelin-1 decreased during heating in controls but not in patients. CONCLUSIONS: These results show that an impaired role of epoxyeicosatrienoic acids contributes, together with an alteration in NO/reactive oxygen species balance and endothelin-1 pathway, to conduit artery endothelial dysfunction in essential hypertension. CLINICAL TRIAL REGISTRATION: https://www.eudract.ema.europa.eu. Unique identifier: RCB2007-A001-10-53.

Changes in carotid arterial wall viscosity and carotid arterial stiffness in type 2 diabetes patients.

BACKGROUND AND AIMS: Changes in arterial wall viscosity (AWW) and stiffness during type 2 diabetes (T2D) have been little investigated. We explored changes in carotid AWV considering change in arterial stiffness and loading conditions, in patients with T2D. METHODS: This cross-sectional, monocentric study compared 19 middle-aged patients with T2D to 30 non-diabetic (ND) controls. The absolute viscosity (WV) was determined as the area of the pressure-lumen cross-sectional area (P-LCSA) loop obtained by carotid tonometry and contralateral echo-tracking. The relative viscosity was determined as the ratio between WV and the elastic energy stored within the arterial wall (WV/WE). Carotid geometry, midwall stress, distensibility and elastic modulus were also compared between groups. RESULTS: T2D patients were older and more frequently had hypertension. Internal diameter, mean central and pulse blood pressure were higher in T2D patients but midwall stress was similar compared to ND controls. WV and WV/WE were higher in T2D patients when compared with ND controls (23 [16-41] vs. 11 [7-18] mm Hg.mm2, p=0.007 and 21% [17-25] vs. 12% [8-17], p < 0.001 respectively) even after adjustment on confounding factors. Carotid arterial stiffness was higher in T2D patients, but after adjustment this difference was only observed for the highest levels of midwall stress. CONCLUSIONS: Carotid AWV and stiffness are increased in T2D patients but only AWV is significantly increased after considering loading conditions. Whether this increase in energy dissipation within the arterial wall contributes to alter cardiovascular coupling in T2D remains to be established.

Adaptation of Arterial Wall Viscosity to the Short-Term Reduction of Heart Rate: Impact of Aging.

Background Changes in arterial wall viscosity, which dissipates the energy stored within the arterial wall, may contribute to the beneficial effect of heart rate (HR) reduction on arterial stiffness and cardiovascular coupling. However, it has never been assessed in humans and could be altered by aging. We evaluated the effect of a selective HR-lowering agent on carotid arterial wall viscosity and the impact of aging on this effect. Methods and Results This randomized, placebo-controlled, double-blind, crossover study performed in 19 healthy volunteers evaluated the effects of ivabradine (5 mg BID, 1-week) on carotid arterial wall viscosity, mechanics, hemodynamics, and cardiovascular coupling. Arterial wall viscosity was evaluated by the area of the hysteresis loop of the pressure-lumen cross-sectional area relationship, representing the energy dissipated (WV), and by the relative viscosity (WV/WE), with WE representing the elastic energy stored. HR reduction by ivabradine increased WV and WE whereas WV/WE remained stable. In middle-aged subjects (n=11), baseline arterial stiffness and cardiovascular coupling were less favorable, and WE was similar but WV and therefore WV/WE were lower than in youth (n=8). HR reduction increased WV/WE in middle-aged but not in young subjects, owing to a larger increase in WV than WE. These results were supported by the age-related linear increase in WV/WE after HR reduction (P=0.009), explained by a linear increase in WV. Conclusion HR reduction increases arterial wall energy dissipation proportionally to the increase in WE, suggesting an adaptive process to bradycardia. This mechanism is altered during aging resulting in a larger than expected energy dissipation, the impact of which should be assessed. Registration URL: https://www.clinicaltrials.gov; Unique identifier: 2015/077/HP. URL: https://www. eudract.ema.europa.eu; Unique identifier: 2015-002060-17.

Determination of Lipoxygenase, CYP450, and Non-Enzymatic Metabolites of Arachidonic Acid in Essential Hypertension and Type 2 Diabetes.

Type 2 diabetes (T2D) and hypertension (HTN) are common risk factors of cardiovascular diseases (CVD) characterized by chronic low-grade systemic inflammation and impaired endothelial function. This study aimed to assess whether levels of non-enzymatic, lipoxygenase (LOX)- and cytochrome P450 (CYP)-derived arachidonic acid (ARA) metabolites, which are known regulators of vascular homeostasis, are affected by HTN and T2D. For this objective, 17 plasma level derivatives of ARA were quantitated by chromatography coupled with mass spectrometry in 44 patients (12 healthy, 8 HTN, 7 T2D, and 17 HTN + T2D). Effects of hyperglycemic and hyperinsulinemic clamps on ARA metabolite levels were assessed in seven healthy subjects. No significant differences in the plasma levels of ARA metabolites were observed for T2D patients compared with healthy volunteers. HTN was associated with an alteration of ARA metabolite correlation patterns with increased 20-, 19-, 15-, and 8-hydroxyeicosatrienoic acid (HETE). A decrease of 20-HETE was also observed during both hyperglycemic and hyperinsulinemic clamps. Additional experiments are needed to assess whether the modulation of HETE metabolites in HTN may be of interest. Furthermore, although not affected by T2D, it remains to investigate whether the decrease of 20-HETE observed during clamps may be related to the regulation of glucose tolerance and insulin signaling.

The effect of camelina oil on vascular function in essential hypertensive patients with metabolic syndrome: a randomized, placebo-controlled, double-blind study.

BACKGROUND: The effects of a dietary supplementation with the vegetable ω-3 α-linolenic acid (ALA) on cardiovascular homeostasis are unclear. In this context, it would be interesting to assess the effects of camelina oil. OBJECTIVE: This study aimed to assess the cardiovascular and metabolic effects of camelina oil in hypertensive patients with metabolic syndrome. METHODS: In a double-blind, placebo-controlled randomized study, treated essential hypertensive patients with metabolic syndrome received, during 6 mo, either cyclodextrin-complexed camelina oil containing ≈ 1.5 g ALA/d (n = 40) or an isocaloric placebo (n = 41), consisting of the same quantity of cyclodextrins and wheat starch. Anthropometric data, plasma lipids, glycemia, insulinemia, creatininemia, TBARs, high-sensitivity C-reactive protein, and n-3, n-6, and n-9 fatty acids in erythrocyte membranes were measured. Peripheral and central blood pressures, arterial stiffness, carotid intima-media thickness, and brachial artery endothelium-dependent flow-mediated dilatation (FMD) and endothelium-independent dilatation were assessed. RESULTS: Compared with placebo, camelina oil increased ALA (mean ± SD: 0 ± 0.04 compared with 0.08 ± 0.06%, P <0.001), its elongation product EPA (0 ± 0.5 compared with 0.16 ± 0.65%, P <0.05), and the n-9 gondoic acid (GA; 0 ± 0.04 compared with 0.08 ± 0.04%, P <0.001). No between-group difference was observed for cardiovascular parameters. However, changes in FMD were associated with the magnitude of changes in EPA (r = 0.26, P = 0.03). Compared with placebo, camelina oil increased fasting glycemia (-0.2 ± 0.6 compared with 0.3 ± 0.5 mmol/L, P <0.001) and HOMA-IR index (-0.8 ± 2.5 compared with 0.5 ± 0.9, P <0.01), without affecting plasma lipids, or inflammatory and oxidative stress markers. Changes in HOMA-IR index were correlated with the magnitude of changes in GA (r = 0.32, P <0.01). Nutritional intake remained similar between groups. CONCLUSION: ALA supplementation with camelina oil did not improve vascular function but adversely affected glucose metabolism in hypertensive patients with metabolic syndrome. Whether this adverse effect on insulin sensitivity is related to GA enrichment, remains to be elucidated.

Evidence for wall shear stress-dependent t-PA release in human conduit arteries: role of endothelial factors and impact of high blood pressure.

Tissue plasminogen activator (t-PA) converts plasminogen into the serine protease plasmin, which in turn degrades fibrin clots. This study assessed whether an increase in shear stress is associated in humans in vivo with the release of t-PA in peripheral conduit arteries, the impact of high blood pressure and the role of NO and CYP450-derived epoxyeicosatrienoic acids (EETs). Local t-PA levels were quantified at baseline and during a sustained increase in radial artery wall shear stress induced by hand skin heating (from 34 to 44 °C) in a total of 25 subjects, among whom 8 were newly diagnosed essential hypertensive patients. The impact of the brachial infusion of NO synthase (L-NMMA) and CYP450 inhibitors (fluconazole) on t-PA release was assessed. The increase in shear stress induced by heating was associated with an increase in local t-PA release (from 3.0 ± 0.5 to 19.2 ± 5.5 ng/min, n = 25, P < 0.01). The magnitude of t-PA release was positively correlated with the increase in shear stress (r = 0.64, P < 0.001) and negatively correlated with mean blood pressure (r = -0.443, P = 0.027). These associations persisted after multiple adjustments for confounding factors. Finally, t-PA release was reduced by L-NMMA and to a larger extent by the combination of L-NMMA and fluconazole without a change in shear stress. The increase in wall shear stress in the peripheral conduit arteries induces a release of t-PA by a mechanism involving NO and EETs. The alteration of this response by high blood pressure may contribute to reducing the fibrinolytic potential and enhancing the risk of arterial thrombosis during exercise.

Preservation of epoxyeicosatrienoic acid bioavailability prevents renal allograft dysfunction and cardiovascular alterations in kidney transplant recipients.

This study addressed the hypothesis that epoxyeicosatrienoic acids (EETs) synthesized by CYP450 and catabolized by soluble epoxide hydrolase (sEH) are involved in the maintenance of renal allograft function, either directly or through modulation of cardiovascular function. The impact of single nucleotide polymorphisms (SNPs) in the sEH gene EPHX2 and CYP450 on renal and vascular function, plasma levels of EETs and peripheral blood monuclear cell sEH activity was assessed in 79 kidney transplant recipients explored at least one year after transplantation. Additional experiments in a mouse model mimicking the ischemia-reperfusion (I/R) injury suffered by the transplanted kidney evaluated the cardiovascular and renal effects of the sEH inhibitor t-AUCB administered in drinking water (10 mg/l) during 28 days after surgery. There was a long-term protective effect of the sEH SNP rs6558004, which increased EET plasma levels, on renal allograft function and a deleterious effect of K55R, which increased sEH activity. Surprisingly, the loss-of-function CYP2C9*3 was associated with a better renal function without affecting EET levels. R287Q SNP, which decreased sEH activity, was protective against vascular dysfunction while CYP2C8*3 and 2C9*2 loss-of-function SNP, altered endothelial function by reducing flow-induced EET release. In I/R mice, sEH inhibition reduced kidney lesions, prevented cardiac fibrosis and dysfunction as well as preserved endothelial function. The preservation of EET bioavailability may prevent allograft dysfunction and improve cardiovascular disease in kidney transplant recipients. Inhibition of sEH appears thus as a novel therapeutic option but its impact on other epoxyfatty acids should be carefully evaluated.

Comparative effects of sirolimus and cyclosporin on conduit arteries endothelial function in kidney recipients.

This study attempted to establish whether a calcineurin inhibitor (CNI)-free immunosuppressant regimen based on sirolimus (SRL) is associated with a preservation of conduit arteries endothelial function in kidney recipients or not. Twenty-nine kidney recipients were randomized to receive since transplantation SRL (n=15) or cyclosporin A (CsA, n=14) associated with mycophenolate mofetil (MMF) and steroids (6months) in a parallel prospective study. Systolic, diastolic blood pressures, glomerular filtration rate (GFR) and radial artery flow-mediated dilatation (FMD) induced by postischaemic hyperaemia were assessed in a blind manner at one (M1) and 7months (M7) after transplantation. Endothelium-independent dilatation was assessed by glyceryl trinitrate spray. There was no difference between the groups for all vascular parameters at M1. At M7, systolic blood pressure was lower (SRL: 119±3 vs. CsA: 138±4mmHg, P<0.05) and FMD was higher in SRL compared with CsA (SRL: 13.1±0.9 vs. CsA: 9.9±0.9%, P<0.05) without any difference for hyperaemia, endothelium-independent dilatation and GFR (SRL: 66.7±1.05 vs. CsA: 67.5±1.22ml/min). Our results demonstrate that a CNI-free regimen based on SRL and MMF prevents conduit artery endothelial dysfunction compared with CsA and MMF in kidney recipients suggesting a beneficial arterial wall effect that may also contribute to the decrease in systolic blood pressure.

Endothelium-dependent adaptation of arterial wall viscosity during blood flow increase is impaired in essential hypertension.

BACKGROUND AND AIMS: Arterial wall viscosity (AWV) is regulated by endothelium-derived NO and epoxyeicosatrienoic acids (EETs) under baseline physiological conditions. Whether these factors regulate AWV during blood flow increase and whether this mechanism is affected in essential hypertensive patients (HT) remain unknown. METHODS: The evolution of radial artery diameter, wall thickness and arterial pressure in response to an increase in flow induced by hand skin heating were measured in 18 untreated HT and 14 normotensive controls (NT) during local infusion of saline and the respective pharmacological inhibitors of NO-synthase and EETs synthesis by cytochrome P450, L-NMMA and/or fluconazole. AWV was estimated by the ratio of the viscous energy dissipated (WV) to the elastic energy stored (WE) obtained from the pressure-diameter relationship. Concomitant changes in operating conditions, which influence the AWV, were taken into account by calculating the midwall stress. RESULTS: Baseline WV and WE were higher in HT than in NT but WV/WE was similar. In saline condition, WV/WE increased in HT during heating but not in NT. In the presence of L-NMMA and/or fluconazole, WV/WE increased during heating in NT. In contrast, these inhibitors did not modify the increase in WV/WE during heating in HT compared to saline. In all conditions, a larger increase in WV than WE was responsible for the increase in WV/WE. CONCLUSIONS: The release of NO and EETs maintains a stable AWV during flow increase and this endothelial adaptive regulation is lost during essential hypertension, which may promote excessive viscous energy dissipation and cardiovascular uncoupling. Restoration of EETs availability with inhibitors of soluble epoxide hydrolase could thus constitute a promising pharmacological approach to restore the endothelial adaptive regulation of AWV.

Fixed combination of perindopril and indapamide at low dose improves endothelial function in essential hypertensive patients after acute administration.

BACKGROUND: Fixed combination of angiotensin-converting enzyme inhibitors (ACEIs) with thiazide-type diuretics at low dose has been used as first-line therapy for the treatment of essential hypertension but their effect on conduit artery endothelial dysfunction remains unknown. METHODS: Thirteen hypertensive patients were assessed after acute administration of a placebo, fixed combination of perindopril-indapamide at low dose: D1 (2 mg/0.625 mg) and twice this dose: D2 (4 mg/1.25 mg), during a double-blind, randomized, crossover study, and were compared with 13 matched controls. Mean arterial pressure (MAP), radial artery diameter (echotracking) and flow (Doppler) were measured during flow-mediated dilatation (FMD) induced by post-ischemic hyperemia (PIH). PIH was characterized by peak flow and duration of hyperemia (t(1/2)). Endothelium-independent dilatation was assessed by trinitrine. RESULTS: In hypertensive patients compared with controls, basal radial artery diameter and flow, peak flow, and trinitrine responses were similar while MAP was increased (115 +/- 3 vs. 87 +/- 2 mm Hg), t(1/2) was decreased (11.1 +/- 1.9 vs. 17.2 +/- 2.2 s), and FMD was altered (radial diameter increase: 203 +/- 14 vs. 304 +/- 15 microm). Compared with placebo, only D2 decreased MAP (placebo: 115 +/- 3; D1: 112 +/- 4; D2: 103 +/- 4 mm Hg) and increased t(1/2) (placebo: 11.1 +/- 1.9; D1: 8.7 +/- 1.5; D2:13.0 +/- 1.9 s). Conversely, D1 and D2 increased FMD (placebo: 203 +/- 14; D1: 218 +/- 22; D2: 227 +/- 23 microm) with no change in basal diameter and flow, peak flow, and trinitrine responses. CONCLUSION: These results demonstrate that a fixed combination of ACEI/diuretic at low dose significantly improves radial artery FMD in hypertensive patients and suggest a direct effect on conduit artery endothelium that may contribute to vascular protection.

Evidence for a Role of Vascular Endothelium in the Control of Arterial Wall Viscosity in Humans.

Arterial wall viscosity (AWV) is a major cause of energy dissipation along the arterial tree. Its determinants remain controversial but an active endothelial regulation has been suggested. Our objective was to assess in humans the physiological role of endothelium-derived nitric oxide (NO), epoxyeicosatrienoic acids and the effect of modulating smooth muscle tone in the regulation of AWV. We simultaneously measured radial artery diameter, wall thickness, and arterial pressure in healthy volunteers during the local infusion of inhibitors of NO-synthase (NG-monomethyl-l-arginine), epoxyeicosatrienoic acids synthesis by cytochrome P450 (fluconazole), the epoxyeicosatrienoic acids cellular targets calcium-activated potassium channels (tetraethylammonium), alone and in combination. AWV was estimated from the relative viscosity expressed as the ratio of the area of the hysteresis loop of the pressure-diameter relationship to the area under the loading phase. Arterial tone was assessed by measuring change in wall stiffness and midwall stress. NG-monomethyl-l-arginine paradoxically reduced relative viscosity (34.9±8.9%-28.9±8.6%). Conversely, relative viscosity was not modified by fluconazole (33.5±15.5%-32.0±13.6%) but increased by tetraethylammonium (31.7±6.6%-35.7±8.0%). This increase was more marked with NG-monomethyl-l-arginine+fluconazole (31.1±10.7%-43.3±13.2%) and NG-monomethyl-l-arginine+tetraethylammonium (29.5±2.3%-41.5±11.1%) compared with inhibitors alone. Sodium nitroprusside decreased AWV (35.4±2.9%-28.7±2.0%). These effects were associated with parallel change in tone but of different magnitude for similar variations in viscosity, suggesting tone-dependent and independent mechanisms. In conclusion, this is the first demonstration that the endothelial factors, NO and epoxyeicosatrienoic acids, regulate AWV in humans and support the role of arterial tone in this regulation. CLINICAL TRIAL REGISTRATION: URL: https://eudract.ema.europa.eu. Unique identifier: RCB2007-A001-10-53.

Alteration in the availability of epoxyeicosatrienoic acids contributes with NO to the development of endothelial dysfunction in conduit arteries during aging.

BACKGROUND AND AIMS: The mechanisms involved in endothelial dysfunction in humans during aging are largely unknown at the level of conduit arteries. We aimed to asses the role of NO and CYP450 epoxygenases-derived epoxyeicosatrienoic acids (EETs) in the regulation of endothelium-dependent flow-mediated dilatation of conduit arteries during aging. METHODS: Radial artery diameter and mean wall shear stress were determined by echotracking coupled with Doppler in 83 subjects (19-71 years old) during a sustained flow increase induced by hand skin heating, with the brachial infusion of saline or NO-synthase and cytochrome P450 epoxygenase inhibitors (L-NNMA and fluconazole respectively). Local blood sampling was performed for the quantification of NO metabolite nitrite and EETs. RESULTS: The magnitude of flow-mediated dilatation was independently and negatively correlated with age, baseline artery diameter and systolic blood pressure, and positively correlated with the increase in shear stress induced by heating. There was an increase in nitrite level during heating until the age of 35-40 years, which declined thereafter. However, the inhibitory effect of L-NMMA on flow-mediated dilatation progressively decreased during aging, demonstrating a decrease in functional NO availability. Moreover, aging progressively reduced the increase in EET level during heating as well as the inhibitory effect of fluconazole on flow-mediated dilatation. CONCLUSIONS: These results show that aging impairs the availability of EETs and NO and epoxyeicosatrienoic acids in peripheral conduit arteries, contributing to the development of endothelial dysfunction.

AT1 receptor blockade prevents the decrease in conduit artery flow-mediated dilatation during NOS inhibition in humans.

Whether AT(1) (angiotenin II type 1) receptor blockade can prevent the decrease in conduit artery FMD (flow-mediated dilatation) during NOS (nitric oxide synthase) inhibition by alternative endothelial pathways has not been explored previously in humans. In 12 healthy subjects, we measured radial artery diameter (echotracking) and flow (Doppler) during FMD induced by sustained reactive hyperaemia during a control period and following NOS inhibition [1.5 mg.min(-1).l(-1) L-NMMA (N(G)-monomethyl-L-arginine)], after a single oral administration of telmisartan (80 mg) or placebo, using a randomized double-blind cross-over design. In six volunteers, we also assessed the roles of prostacyclin and EDHF (endothelium-derived hyperpolarizing factor) during radial FMD after AT(1) receptor blockade by oral administration of aspirin (500 mg) alone, aspirin+L-NMMA or aspirin+L-NMMA+fluconazole (a cytochrome epoxygenases inhibitor; 0.37 mg.min(-1).l(-1)). Telmisartan did not affect radial artery FMD in the control period (10.9+/-0.6% with placebo compared with 9.9+/-0.7% with telmisartan), but prevented its decrease after L-NMMA (9.3+/-0.8% with placebo compared with 12.6+/-1.2% with telmisartan; P<0.05) with no modification in baseline parameters, hyperaemia and radial artery endothelium-independent dilatation to sodium nitroprusside. Moreover, in telmisartan-treated subjects, radial artery FMD, compared with control (9.0+/-1.0%), was not modified by aspirin alone (9.4+/-0.7%) or associated with L-NMMA (9.5+/-0.5%), but was reduced by the combination of aspirin, L-NMMA and fluconazole (7.5+/-0.6%; P<0.05). These results demonstrate that AT(1) receptor blockade prevents the decrease in conduit artery FMD during NOS inhibition in humans, suggesting the development of a compensatory endothelial mechanism. This mechanism appears to be independent of prostacyclin and could possibly be related to an EDHF release.

Physiological role of endothelin-1 in flow-mediated vasodilatation in humans and impact of cardiovascular risk factors.

OBJECTIVES: The current study addressed the hypothesis that the local decrease in endothelin-1 (ET-1) bioavailability during sustained flow increases contributes to endothelium-dependent, flow-mediated dilatation (FMD) of conduit arteries and is altered in presence of cardiovascular risk factors. METHODS AND RESULTS: In nine young healthy individuals, the decrease in local ET-1 plasma levels and radial artery FMD in response to hand skin heating (from 34 to 44 °C) was not affected by endothelin type A (ETA) receptor blockade, achieved using the brachial infusion of BQ-123 (100 nmol/min per l of forearm), as compared with physiological saline (0.9% NaCl) infusion. In contrast, endothelin type B (ETB) receptor blockade with BQ-788 (10 nmol/min per l) suppressed the decrease in plasma ET-1 during heating and reduced FMD, without altering nitric oxide release. The coinfusion of BQ-123 did not affect the inhibitory effect of ETB receptor blockade on the decrease in ET-1 plasma levels during heating but prevented the reduction in FMD. Basal radial artery parameters, systemic hemodynamics, and endothelium-independent dilatation to glyceryl trinitrate were not modified by ETA and/or ETB blockade. In a general population of 40 participants without treatment or major cardiovascular diseases, including the nine healthy individuals, the reduction in endothelin-1 level during heating was correlated with FMD (r = -0.55, P < 0.001) and decreased with increased age (r = 0.49, P = 0.001), mean arterial blood pressure (r = 0.48, P = 0.002), and total cholesterol level (r = 0.37, P = 0.024). CONCLUSION: The uptake of endothelin-1 by ETB receptors contributes to conduit artery FMD, preventing its vasoconstrictor action mediated by ETA receptors. The alteration of this mechanism by cardiovascular risk factors may contribute to endothelial dysfunction.

Role of Toll-like receptors 2 and 4 in mediating endothelial dysfunction and arterial remodeling in primary arterial antiphospholipid syndrome.

OBJECTIVE: To assess the role of Toll-like receptors (TLRs) in antiphospholipid antibody (aPL)-mediated vascular abnormalities in patients with primary arterial antiphospholipid syndrome (APS). METHODS: Forty-eight subjects participated in the study. Arterial function and structure and TLR pathway activation were determined in patients with primary arterial APS and matched controls. The pathogenic effects of aPL isolated from patients were assessed in wild-type (WT) and TLR-knockout mice. RESULTS: APS patients had endothelial dysfunction, arterial stiffening, and hypertrophy, as evidenced by decreased brachial artery endothelium-dependent flow-mediated dilation (FMD) and increased aortic pulse wave velocity and carotid intima-media thickness (IMT), as compared with controls. Plasma samples from APS patients revealed decreased nitric oxide (NO) availability and a pro-oxidative, proinflammatory, and prothrombotic state illustrated by a decrease in nitrite and an increase in lipid peroxidation, tumor necrosis factor α levels, and tissue factor (TF) levels. Furthermore, TLR pathway activation was found in APS patients with increased TLR-2 and TLR-4 messenger RNA expression and increased protein levels of the activated TLR transduction protein interleukin-1 receptor-associated kinase 1 in peripheral blood mononuclear cells. Moreover, agonist-stimulated cell-surface expression of TLR-2 and TLR-4 in circulating monocytes was higher in APS patients than in controls. These changes were positively associated with IMT and negatively associated with FMD. Finally, aPL injection decreased mesenteric endothelium-dependent relaxation and increased TF expression in WT mice but not in TLR-2- or TLR-4-knockout mice. CONCLUSION: This translational study supports the notion that TLR-2 and TLR-4 play a role in mediating vascular abnormalities in patients with primary arterial APS. TLRs thus constitute a promising pharmacologic target for preventing cardiovascular complications in APS.