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.

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.

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.

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.

Pharmacokinetic modeling of morphine and its glucuronides: Comparison of nebulization versus intravenous route in healthy volunteers.

Intravenous (i.v.) morphine is a safe, robust, and recommended treatment for severe pain using the titration principle. Despite its high efficacy, it is impacted by organizational constraints related to venous access. Nebulized (NEB) morphine may represent an alternative for titration but pharmacokinetic (PK) properties of short nebulization using routine devices need evaluation. Twenty-seven healthy volunteers were included to receive NEB or i.v. morphine administration using increasing amounts according to Dixon's reference method. Plasma morphine, morphine-3-glucuronide (M3G), and morphine-6-glucuronide (M6G) were quantified. PK modeling and simulations were performed using Monolix. Dixon's method exhibited a significantly higher morphine dose regimen in the NEB group versus the i.v. group (6.2 [5.3-7.1] vs. 3.0 [2.0-4.0] mg, p < 0.001). Morphine, M3G, and M6G dose-normalized exposure were significantly lower in the NEB group versus the i.v. group: morphine (19 [13-23] vs. 1044 [702-1266] µg min/L, p < 0.001), M3G (245 [162-287] vs. 3752 [2487-5165] µg min/L, p < 0.001) and M6G (28 [21-43] vs. 466 [370-723] µg min/L, p < 0.001). The model that best fitted the data consisted in a transit compartment for morphine absorption, three compartments for morphine distribution followed by multiple transit compartments (8.2 and 57.5-min transit time for M3G and M6G, respectively) and a first order elimination for M3G and M6G. Morphine bioavailability in the NEB group was 3.5% using the i.v. group as reference. Administration route and sex significantly influenced morphine and metabolite PKs. This work aimed to evaluate the PKs of NEB morphine compared with the i.v. route. Despite a bioavailability to improve, NEB morphine administration using a routine device is suitable to plan morphine titration.

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.

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.

Deleterious impact of a generic temozolomide formulation compared with brand-name product on the kinetic of platelet concentration and survival in newly diagnosed glioblastoma.

Chemo-induced thrombocytopenia is a limiting toxicity among patients receiving temozolomide (TMZ) as first-line treatment for glioblastoma. We aimed to compare early platelet concentration kinetics, hematological safety profile, and impact on survival following the initiation of either the brand-name or a generic TMZ formulation. A retrospective trial was conducted in patients suffering from newly diagnosed glioblastoma. Patients were treated with TMZ at 75 mg/m2 per day during six weeks, concomitantly with radiotherapy. Platelet concentration was collected each week. Primary endpoint was to perform a linear mixed-effect model of platelet concentration kinetic over weeks. A total of 147 patients were included as follows: 96 received the brand-name TMZ, and 51 received a generic TMZ formulation. Exposition to the generic was a significant variable that negatively influenced the platelet kinetics in the radiotherapy and concomitant TMZ phase, P = 0.02. Grade ≥3 chemo-induced thrombocytopenia was more frequent in the generic group: 19.6% [95% CI 8.7-30.5%] vs 3.1% [0-6.6%], P = 0.001. Exposition to the generic formulation of TMZ led to increase early treatment discontinuation due to TMZ-induced thrombocytopenia and was a worsening independent prognostic factor on overall survival: adjusted HR 1.83 [1.21-2.8], P = 0.031. These data suggest that exposition to a generic formulation of TMZ vs the brand-name product is associated with higher early platelet decrease leading to clinically relevant impacts on treatment schedule in glioblastoma. Further prospective trials are needed to confirm these results.

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.

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.

Hydroxychloroquine reverses the prothrombotic state in a mouse model of antiphospholipid syndrome: Role of reduced inflammation and endothelial dysfunction.

Antiphospholipid antibodies (aPL) promote endothelial dysfunction, inflammation and procoagulant state. We investigated the effect of hydroxychloroquine (HCQ) on prothrombotic state and endothelial function in mice and in human aortic endothelial cells (HAEC). Human aPL were injected to C57BL/6 mice treated or not with HCQ. Vascular endothelial function and eNOS were assessed in isolated mesenteric arteries. Thrombosis was assessed both in vitro by measuring thrombin generation time (TGT) and tissue factor (TF) expression and in vivo by the measurement of the time to occlusion in carotid and the total thrombosis area in mesenteric arteries. TGT, TF, and VCAM1 expression were evaluated in HAEC. aPL increased VCAM-1 expression and reduced endothelium dependent relaxation to acetylcholine. In parallel, aPL shortened the time to occlusion and extended thrombus area in mice. This was associated with an overexpression of TF and an increased TGT in mice and in HAEC. HCQ reduced clot formation as well as TGT, and improved endothelial-dependent relaxations. Finally, HCQ increased the p-eNOS/eNOS ratio. This study provides new evidence that HCQ improves procoagulant status and vascular function in APS by modulating eNOS, leading to an improvement in the production of NO.

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.

Possible role of CYP2B6 genetic polymorphisms in ifosfamide-induced encephalopathy: report of three cases.

Ifosfamide (IFA) is a potent alkylating antitumoral agent, but its use is limited by neurological side effects. IFA is a racemic mixture of two enantiomeric forms, R-IFA and S-IFA with a stereoselective metabolism by CYP3A4 and CYP2B6, leading either to bioactive or to toxic pathways. In three consecutive cases of pediatric patients who exhibited IFA-induced encephalopathy (IIE), genotyping of clinically relevant single-nucleotide polymorphisms associated with decreased CYP3A4 and CYP2B6 activities was performed. Genetic investigations revealed the presence of CYP2B6 rs4803419 (C>T) in one patient while the two others carried the CYP2B6*6 allelic variant. All patients carried CYP3A4 wild-type genotype (CYP3A4*1/*1). Because CYP2B6-deficient alleles may be responsible for an increased conversion of S-IFA into neurotoxic metabolites, screening for CYP2B6 polymorphisms may help to avoid IIE and improve clinical outcomes.

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.

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.

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.

Infliximab improves endothelial dysfunction in a mouse model of antiphospholipid syndrome: Role of reduced oxidative stress.

Antiphospholipid syndrome (APS), induces endothelial dysfunction, oxidative stress and systemic inflammation that may be mediated by TNFα. Thus, we investigated the possible protective effect of the anti-TNFα antibody infliximab (5µg/g) on endothelial function in a mouse APS model (induced by injection of purified human anti-ß2GP1-IgG). Seven days after anti-ß2GPI-IgG injection, we observed an increase in plasma sVCAM-1 and sE-selectin levels and in aortic mRNA expression of VCAM-1 and E-selectin. This was associated with a decreased endothelium-dependent relaxation of isolated mesenteric arteries to acetylcholine, together with decreased mesenteric eNOS mRNA expression and increased eNOS uncoupling, accompanied by increased iNOS and gp91phox mRNA and increased left ventricular GSH/GSSH ratio. Infliximab significantly improved the NO-mediated relaxing responses to acetylcholine, and induced a decrease in iNOS and gp91phox mRNA expression. The õpro-adhesive and pro-coagulant phenotypes induced by the anti-ß2GP1-IgG were also reversed. This study provides the first evidence that TNFα antagonism improves endothelial dysfunction in APS and suggests that endothelial dysfunction is mediated by TNFα and oxidative stress. Therefore, infliximab may be of special relevance in clinical practice.

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.