Publications by authors named "Julie Favre"

28 Publications

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

Sci Rep 2021 Feb 12;11(1):3739. Epub 2021 Feb 12.

Department of Pharmacology, Rouen University Hospital, 76000, Rouen, France.

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.
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http://dx.doi.org/10.1038/s41598-021-83274-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7881112PMC
February 2021

Endothelial S1P Signaling Counteracts Infarct Expansion in Ischemic Stroke.

Circ Res 2021 Feb 2;128(3):363-382. Epub 2020 Dec 2.

Vascular Biology Program, Boston Children's Hospital, MA (T.H.).

Rationale: Cerebrovascular function is critical for brain health, and endogenous vascular protective pathways may provide therapeutic targets for neurological disorders. S1P (Sphingosine 1-phosphate) signaling coordinates vascular functions in other organs, and S1P (S1P receptor-1) modulators including fingolimod show promise for the treatment of ischemic and hemorrhagic stroke. However, S1P also coordinates lymphocyte trafficking, and lymphocytes are currently viewed as the principal therapeutic target for S1P modulation in stroke.

Objective: To address roles and mechanisms of engagement of endothelial cell S1P in the naive and ischemic brain and its potential as a target for cerebrovascular therapy.

Methods And Results: Using spatial modulation of S1P provision and signaling, we demonstrate a critical vascular protective role for endothelial S1P in the mouse brain. With an S1P signaling reporter, we reveal that abluminal polarization shields S1P from circulating endogenous and synthetic ligands after maturation of the blood-neural barrier, restricting homeostatic signaling to a subset of arteriolar endothelial cells. S1P signaling sustains hallmark endothelial functions in the naive brain and expands during ischemia by engagement of cell-autonomous S1P provision. Disrupting this pathway by endothelial cell-selective deficiency in S1P production, export, or the S1P receptor substantially exacerbates brain injury in permanent and transient models of ischemic stroke. By contrast, profound lymphopenia induced by loss of lymphocyte S1P provides modest protection only in the context of reperfusion. In the ischemic brain, endothelial cell S1P supports blood-brain barrier function, microvascular patency, and the rerouting of blood to hypoperfused brain tissue through collateral anastomoses. Boosting these functions by supplemental pharmacological engagement of the endothelial receptor pool with a blood-brain barrier penetrating S1P-selective agonist can further reduce cortical infarct expansion in a therapeutically relevant time frame and independent of reperfusion.

Conclusions: This study provides genetic evidence to support a pivotal role for the endothelium in maintaining perfusion and microvascular patency in the ischemic penumbra that is coordinated by S1P signaling and can be harnessed for neuroprotection with blood-brain barrier-penetrating S1P agonists.
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http://dx.doi.org/10.1161/CIRCRESAHA.120.316711DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7874503PMC
February 2021

Mutation of Arginine 264 on ERα (Estrogen Receptor Alpha) Selectively Abrogates the Rapid Signaling of Estradiol in the Endothelium Without Altering Fertility.

Arterioscler Thromb Vasc Biol 2020 09 9;40(9):2143-2158. Epub 2020 Jul 9.

From the INSERM-UPS UMR U1048, Institut des Maladies Métaboliques et Cardiovasculaires (M.A., C.F., R.Z., M.B., R.S., T.A., F.B., P.G., H.L., C.F., J.-F.A., F.L.), Université de Toulouse, France.

Objective: ERα (estrogen receptor alpha) exerts nuclear genomic actions and also rapid membrane-initiated steroid signaling. The mutation of the cysteine 451 into alanine in vivo has recently revealed the key role of this ERα palmitoylation site on some vasculoprotective actions of 17β-estradiol (E2) and fertility. Here, we studied the in vivo role of the arginine 260 of ERα which has also been described to be involved in its E2-induced rapid signaling with PI-3K (phosphoinositide 3-kinase) as well as G protein in cultured cell lines. Approach and Results: We generated a mouse model harboring a point mutation of the murine counterpart of this arginine into alanine (R264A-ERα). In contrast to the , the females are fertile with standard hormonal serum levels and normal control of hypothalamus-pituitary ovarian axis. Although R264A-ERα protein abundance was normal, the well-described membrane ERα-dependent actions of estradiol, such as the rapid dilation of mesenteric arteries and the acceleration of endothelial repair of carotid, were abrogated in mice. In striking contrast, E2-regulated gene expression was highly preserved in the uterus and the aorta, revealing intact nuclear/genomic actions in response to E2. Consistently, 2 recognized nuclear ERα-dependent actions of E2, namely atheroma prevention and flow-mediated arterial remodeling were totally preserved.

Conclusions: These data underline the exquisite role of arginine 264 of ERα for endothelial membrane-initiated steroid signaling effects of E2 but not for nuclear/genomic actions. This provides the first model of fertile mouse with no overt endocrine abnormalities with specific loss-of-function of rapid ERα signaling in vascular functions.
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http://dx.doi.org/10.1161/ATVBAHA.120.314159DOI Listing
September 2020

Nuclear Activation Function 2 Estrogen Receptor α Attenuates Arterial and Renal Alterations Due to Aging and Hypertension in Female Mice.

J Am Heart Assoc 2020 03 27;9(5):e013895. Epub 2020 Feb 27.

MITOVASC Institute and CARFI Facility INSERM U1083 CNRS UMR 6015 Angers University Angers France.

Background The cardiovascular protective effects of estrogens in premenopausal women depend mainly on estrogen receptor α (ERα). ERα activates nuclear gene transcription regulation and membrane-initiated signaling. The latter plays a key role in estrogen-dependent activation of endothelial NO synthase. The goal of the present work was to determine the respective roles of the 2 ERα activities in endothelial function and cardiac and kidney damage in young and old female mice with hypertension, which is a major risk factor in postmenopausal women. Methods and Results Five- and 18-month-old female mice lacking either ERα (ERα), the nuclear activating function AF2 of ERα (AF2°), or membrane-located ERα (C451A) were treated with angiotensin II (0.5 mg/kg per day) for 1 month. Systolic blood pressure, left ventricle weight, vascular reactivity, and kidney function were then assessed. Angiotensin II increased systolic blood pressure, ventricle weight, and vascular contractility in ERα and AF2° mice more than in wild-type and C451A mice, independent of age. In both the aorta and mesenteric resistance arteries, angiotensin II and aging reduced endothelium-dependent relaxation in all groups, but this effect was more pronounced in ERα and AF2° than in the wild-type and C451A mice. Kidney inflammation and oxidative stress, as well as blood urea and creatinine levels, were also more pronounced in old hypertensive ERα and AF2° than in old hypertensive wild-type and C451A mice. Conclusions The nuclear ERα-AF2 dependent function attenuates angiotensin II-dependent hypertension and protects target organs in aging mice, whereas membrane ERα signaling does not seem to play a role.
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http://dx.doi.org/10.1161/JAHA.119.013895DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7335584PMC
March 2020

Evidence of Cardiovascular Calcification and Fibrosis in Pseudoxanthoma Elasticum Mouse Models Subjected to DOCA-Salt Hypertension.

Sci Rep 2019 11 8;9(1):16327. Epub 2019 Nov 8.

MitoVasc Institute, UMR CNRS 6015 - INSERM U1083, Angers University, Angers, France.

Pseudoxanthoma Elasticum (PXE) is a rare disorder characterized by fragmentation and progressive calcification of elastic fibres in connective tissues. Although arterial hypertension (AHT) has been reported in PXE patients, its impact on pathological manifestations has as yet been unexplored. We investigated the consequences of experimental AHT on Abcc6-/- PXE mouse models. Experimental AHT was induced by deoxycorticosterone acetate (DOCA-salt) in uni-nephrectomised mice. Blood pressure (BP) and vascular reactivity were monitored using tail-cuff plethysmography and myography respectively. Calcium content and fibrosis were assessed using colorimetry, Von Kossa and Sirius red staining respectively. The gene expression implicated in vascular biology was measured using quantitative polymerase chain reaction. DOCA-salt induced a matching rise in BP in Abcc6-/- and WT mice. Aortic ring contraction and relaxation in vitro were comparable. Calcium accumulated in the hearts of hypertensive Abcc6-/- mice along with significant fibrosis in the myocardium and aorta by contrast with the WT mice. In hypertensive Abcc6-/- mouse aortas, these results were corroborated by gene expression patterns favouring calcification, fibrosis and extracellular matrix remodelling. Abcc6 loss-of-function is associated with greater cardiovascular calcification and fibrosis in mice subjected to DOCA-Salt hypertension. These results suggest likely cardiovascular deterioration in PXE patients with AHT, necessitating diligent BP monitoring.
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http://dx.doi.org/10.1038/s41598-019-52808-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6841718PMC
November 2019

Myogenic vasoconstriction requires G/G and LARG to maintain local and systemic vascular resistance.

Elife 2019 09 24;8. Epub 2019 Sep 24.

Department of Pharmacology, Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany.

Myogenic vasoconstriction is an autoregulatory function of small arteries. Recently, G-protein-coupled receptors have been involved in myogenic vasoconstriction, but the downstream signalling mechanisms and the in-vivo-function of this myogenic autoregulation are poorly understood. Here, we show that small arteries from mice with smooth muscle-specific loss of G/G or the Rho guanine nucleotide exchange factor ARHGEF12 have lost myogenic vasoconstriction. This defect was accompanied by loss of RhoA activation, while vessels showed normal increases in intracellular [Ca]. In the absence of myogenic vasoconstriction, perfusion of peripheral organs was increased, systemic vascular resistance was reduced and cardiac output and left ventricular mass were increased. In addition, animals with defective myogenic vasoconstriction showed aggravated hypotension in response to endotoxin. We conclude that G/G- and Rho-mediated signaling plays a key role in myogenic vasoconstriction and that myogenic tone is required to maintain local and systemic vascular resistance under physiological and pathological condition.
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http://dx.doi.org/10.7554/eLife.49374DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6777979PMC
September 2019

Predominant Role of Nuclear Versus Membrane Estrogen Receptor α in Arterial Protection: Implications for Estrogen Receptor α Modulation in Cardiovascular Prevention/Safety.

J Am Heart Assoc 2018 06 29;7(13). Epub 2018 Jun 29.

Institut des Maladies Métaboliques et Cardiovasculaires (I2MC), Institut National de la Santé et de la Recherche Médicale (INSERM) U 1048, University of Toulouse 3, France.

Background: Although estrogen receptor α (ERα) acts primarily as a transcription factor, it can also elicit membrane-initiated steroid signaling. Pharmacological tools and transgenic mouse models previously highlighted the key role of ERα membrane-initiated steroid signaling in 2 actions of estrogens in the endothelium: increase in NO production and acceleration of reendothelialization.

Methods And Results: Using mice with ERα mutated at cysteine 451 (ERaC451A), recognized as the key palmitoylation site required for ERα plasma membrane location, and mice with disruption of nuclear actions because of inactivation of activation function 2 (ERaAF20 = ERaAF2°), we sought to fully characterize the respective roles of nuclear membrane-initiated steroid signaling in the arterial protection conferred by ERα. ERaC451A mice were fully responsive to estrogens to prevent atheroma and angiotensin II-induced hypertension as well as to allow flow-mediated arteriolar remodeling. By contrast, ERαAF20 mice were unresponsive to estrogens for these beneficial vascular effects. Accordingly, selective activation of nuclear ERα with estetrol was able to prevent hypertension and to restore flow-mediated arteriolar remodeling.

Conclusions: Altogether, these results reveal an unexpected prominent role of nuclear ERα in the vasculoprotective action of estrogens with major implications in medicine, particularly for selective nuclear ERα agonist, such as estetrol, which is currently under development as a new oral contraceptive and for hormone replacement therapy in menopausal women.
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http://dx.doi.org/10.1161/JAHA.118.008950DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6064913PMC
June 2018

GPR68 Senses Flow and Is Essential for Vascular Physiology.

Cell 2018 04;173(3):762-775.e16

Doris Neuroscience Center, Howard Hughes Medical Institute, the Scripps Research Institute, La Jolla, CA 92037, USA. Electronic address:

Mechanotransduction plays a crucial role in vascular biology. One example of this is the local regulation of vascular resistance via flow-mediated dilation (FMD). Impairment of this process is a hallmark of endothelial dysfunction and a precursor to a wide array of vascular diseases, such as hypertension and atherosclerosis. Yet the molecules responsible for sensing flow (shear stress) within endothelial cells remain largely unknown. We designed a 384-well screening system that applies shear stress on cultured cells. We identified a mechanosensitive cell line that exhibits shear stress-activated calcium transients, screened a focused RNAi library, and identified GPR68 as necessary and sufficient for shear stress responses. GPR68 is expressed in endothelial cells of small-diameter (resistance) arteries. Importantly, Gpr68-deficient mice display markedly impaired acute FMD and chronic flow-mediated outward remodeling in mesenteric arterioles. Therefore, GPR68 is an essential flow sensor in arteriolar endothelium and is a critical signaling component in cardiovascular pathophysiology.
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http://dx.doi.org/10.1016/j.cell.2018.03.076DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5951615PMC
April 2018

Protein tyrosine phosphatase 1B inactivation limits aging-associated heart failure in mice.

Am J Physiol Heart Circ Physiol 2018 06 23;314(6):H1279-H1288. Epub 2018 Mar 23.

Normandie University UNIROUEN, Institut National de la Santé et de la Recherche Médicale U1096 , Rouen , France.

We have previously shown that protein tyrosine phosphatase 1B (PTP1B) inactivation in mice [PTP1B-deficient (PTP1B) mice] improves left ventricular (LV) angiogenesis, perfusion, remodeling, and function and limits endothelial dysfunction after myocardial infarction. However, whether PTP1B inactivation slows aging-associated cardiovascular dysfunction remains unknown. Wild-type (WT) and PTP1B mice were allowed to age until 18 mo. Compared with old WT mice, in which aging increased the LV mRNA expression of PTP1B, old PTP1B mice had 1) reduced cardiac hypertrophy with decreased LV mRNA levels of hypertrophic markers and atrial and brain natriuretic peptides, 2) lower LV fibrosis (collagen: 16 ± 3% in WT mice and 5 ± 3% in PTP1B mice, P < 0.001) with decreased mRNA levels of transforming growth-factor-β and matrix metalloproteinase-2, and 3) higher LV capillary density and lower LV mRNA level of hypoxic inducible factor-1α, which was associated over time with a higher rate of proangiogenic M2 type macrophages and a stable LV mRNA level of VEGF receptor-2. Echocardiography revealed an age-dependent LV increase in end-diastolic volume in WT mice together with alterations of fractional shortening and diastole (transmitral Doppler E-to-A wave ratio). Invasive hemodynamics showed better LV systolic contractility and better diastolic compliance in old PTP1B mice (LV end-systolic pressure-volume relation: 13.9 ± 0.9 in WT mice and 18.4 ± 1.6 in PTP1B mice; LV end-diastolic pressure-volume relation: 5.1 ± 0.8 mmHg/relative volume unit in WT mice and 1.2 ± 0.3 mmHg/relative volume unit in PTP1B mice, P < 0.05). In addition, old PTP1B mice displayed a reduced amount of LV reactive oxygen species. Finally, in isolated resistance mesenteric arteries, PTP1B inactivation reduced aging-associated endothelial dysfunction (flow-mediated dilatation: -0.4 ± 2.1% in WT mice and 8.2 ± 2.8% in PTP1B mice, P < 0.05). We conclude that PTP1B inactivation slows aging-associated LV remodeling and dysfunction and reduces endothelial dysfunction in mesenteric arteries. NEW & NOTEWORTHY The present study shows that protein tyrosine phosphatase 1B inactivation in aged mice improves left ventricular systolic and diastolic function associated with reduced adverse cardiac remodeling (hypertrophy, fibrosis, and capillary rarefaction) and limits vascular endothelial dysfunction. This suggests that protein tyrosine phosphatase 1B inhibition could be an interesting treatment approach in age-related cardiovascular dysfunction.
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http://dx.doi.org/10.1152/ajpheart.00049.2017DOI Listing
June 2018

Loss of vascular expression of nucleoside triphosphate diphosphohydrolase-1/CD39 in hypertension.

Purinergic Signal 2018 03 13;14(1):73-82. Epub 2017 Dec 13.

MITOVASC Institute - UMR CNRS 6015 INSERM U1083 University of Angers, Angers, France.

Ectonucleoside triphosphate diphosphohydrolase-1, the major vascular/immune ectonucleotidase, exerts anti-thrombotic and immunomodulatory actions by hydrolyzing extracellular nucleotides (danger signals). Hypertension is characterized by vascular wall remodeling, endothelial dysfunction, and immune infiltration. Here our aim was to investigate the impact of arterial hypertension on CD39 expression and activity in mice. Arterial expression of CD39 was determined by reverse transcription quantitative real-time PCR in experimental models of hypertension, including angiotensin II (AngII)-treated mice (1 mg/kg/day, 21 days), deoxycorticosterone acetate-salt mice (1% salt and uninephrectomy, 21 days), and spontaneously hypertensive rats. A decrease in CD39 expression occurred in the resistance and conductance arteries of hypertensive animals with no effect on lymphoid organs. In AngII-treated mice, a decrease in CD39 protein levels (Western blot) was corroborated by reduced arterial nucleotidase activity, as evaluated by fluorescent (etheno)-ADP hydrolysis. Moreover, serum-soluble ADPase activity, supported by CD39, was significantly decreased in AngII-treated mice. Experiments were conducted in vitro on vascular cells to determine the elements underlying this downregulation. We found that CD39 transcription was reduced by proinflammatory cytokines interleukin (IL)-1β and tumor necrosis factor alpha on vascular smooth muscle cells and by IL-6 and anti-inflammatory and profibrotic cytokine transforming growth factor beta 1 on endothelial cells. In addition, CD39 expression was downregulated by mechanical stretch on vascular cells. Arterial expression and activity of CD39 were decreased in hypertension as a result of both a proinflammatory environment and mechanical strain exerted on vascular cells. Reduced ectonucleotidase activity may alter the vascular condition, thus enhancing arterial damage, remodeling, or thrombotic events.
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http://dx.doi.org/10.1007/s11302-017-9597-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5842157PMC
March 2018

Modulating putative endothelial progenitor cells for the treatment of endothelial dysfunction and cardiovascular complications in diabetes.

Pharmacol Ther 2017 02 20;170:98-115. Epub 2016 Oct 20.

Department of Pharmacology, Rouen University Hospital, Rouen, France; Institut National de la Santé et de la Recherche Médicale (INSERM) U1096, Rouen, France; University of Rouen, Institute for Research and Innovation in Biomedicine, Rouen, France. Electronic address:

Diabetes induces a decrease in the number and function of different pro-angiogenic cell types generically designated as putative endothelial progenitor cells (EPC), which encompasses cells from myeloid origin that act in a paracrine fashion to promote angiogenesis and putative "true" EPC that contribute to endothelial replacement. This not only compromises neovasculogenesis in ischemic tissues but also impairs, at an early stage, the reendotheliziation process at sites of injury, contributing to the development of endothelial dysfunction and cardiovascular complications. Hyperglycemia, insulin resistance and dyslipidemia promote putative EPC dysregulation by affecting the SDF-1/CXCR-4 and NO pathways and the p53/SIRT1/p66Shc axis that contribute to their mobilization, migration, homing and vasculogenic properties. To optimize the clinical management of patients with hypoglycemic agents, statins and renin-angiotensin system inhibitors, which display pleiotropic effects on putative EPC, is a first step to improve their number and angiogenic potential but specific strategies are needed. Among them, mobilizing therapies based on G-CSF, erythropoietin or CXCR-4 antagonism have been developed to increase putative EPC number to treat ischemic diseases with or without prior cell isolation and transplantation. Growth factors, genetic and pharmacological strategies are also evaluated to improve ex vivo cultured EPC function before transplantation. Moreover, pharmacological agents increasing in vivo the bioavailability of NO and other endothelial factors demonstrated beneficial effects on neovascularization in diabetic ischemic models but their effects on endothelial dysfunction remain poorly evaluated. More experiments are warranted to develop orally available drugs and specific agents targeting p66Shc to reverse putative EPC dysfunction in the expected goal of preventing endothelial dysfunction and diabetic cardiovascular complications.
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http://dx.doi.org/10.1016/j.pharmthera.2016.10.014DOI Listing
February 2017

Vascular Smooth Muscle Mineralocorticoid Receptor Contributes to Coronary and Left Ventricular Dysfunction After Myocardial Infarction.

Hypertension 2016 Apr 22;67(4):717-23. Epub 2016 Feb 22.

From the Institut National de la Santé et de la Recherche Médicale (Inserm) U1096, Rouen, France (A.G., N.H., J.F., L.N., J.-P.H., M.B., C.T., V.R., P.M., A.O.-P.); Institute for Research and Innovative Biomedicine (A.G., N.H., J.F., L.N., J.-P.H., M.B., C.T., V.R., P.M., A.O.-P.), UFR Médecine-Pharmacie (A.G., N.H., J.F., L.N., J.-P.H., M.B., C.T., V.R., P.M., A.O.-P.), and Plateau d'Imagerie Cardio-Thoracique de l'Université de Rouen (PICTUR) (L.N., C.T., P.M.), Normandy-University, Rouen, France; Inserm U1138, Cordeliers Institute, Paris VI-University, Paris, France (G.G., F.J.); and Cardiology Research, Bayer-Pharmaceuticals, Wuppertal, Germany (P.K.).

Mineralocorticoid receptor (MR) antagonists slow down the progression of heart failure after myocardial infarction (MI), but the cell-specific role of MR in these benefits is unclear. In this study, the role of MR expressed in vascular smooth muscle cells (VSMCs) was investigated. Two months after coronary artery ligation causing MI, mice with VSMC-specific MR deletion (MI-MR(SMKO)) and mice treated with the MR antagonist finerenone (MI-fine) had improved left ventricular compliance and elastance when compared with infarcted control mice (MI-CTL), as well as reduced interstitial fibrosis. Importantly, the coronary reserve assessed by magnetic resonance imaging was preserved (difference in myocardial perfusion before and after induction of vasodilatation, mL mg(-1) min(-1): MI-CTL: 1.1 ± 0.5, nonsignificant; MI-MR(SMKO): 4.6 ± 1.6 [P<0.05]; MI-fine: 3.6 ± 0.7 [P<0.01]). The endothelial function, tested on isolated septal coronary arteries by analyzing the acetylcholine-induced nitric oxide-dependent relaxation, was also improved by MR deletion in VSMCs or by finerenone treatment (relaxation %: MI-CTL: 36 ± 5, MI-MR(SMKO): 54 ± 3, and MI-fine: 76 ± 4; P<0.05). Such impairment of the coronary endothelial function on MI involved an oxidative stress that was reduced when MR was deleted in VSMCs or by finerenone treatment. Moreover, short-term incubation of coronary arteries isolated from noninfarcted animals with low-dose angiotensin-II (10(-9) mol/L) induced oxidative stress and impaired acetylcholine-induced relaxation in CTL but neither in MR(SMKO) nor in mice pretreated with finerenone. In conclusion, deletion of MR in VSMCs improved left ventricular dysfunction after MI, likely through maintenance of the coronary reserve and improvement of coronary endothelial function. MR blockage by finerenone had similar effects.
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http://dx.doi.org/10.1161/HYPERTENSIONAHA.115.06709DOI Listing
April 2016

IFN-β affects the angiogenic potential of circulating angiogenic cells by activating calpain 1.

Am J Physiol Heart Circ Physiol 2015 Nov 2;309(10):H1667-78. Epub 2015 Oct 2.

Department of Molecular Cell Biology and Immunology, VU University Medical Center, Amsterdam, The Netherlands;

Circulating angiogenic cells (CACs) are monocyte-derived cells with endothelial characteristics, which contribute to both angiogenesis and arteriogenesis in a paracrine way. Interferon-β (IFN-β) is known to inhibit these divergent processes in animals and patients. We hypothesized that IFN-β might act by affecting the differentiation and function of CACs. CACs were cultured from peripheral blood mononuclear cells and phenotypically characterized by surface expression of monocytic and endothelial markers. IFN-β significantly reduced the number of CACs by 18-64%. Apoptosis was not induced by IFN-β, neither in mononuclear cells during differentiation, nor after maturation to CACs. Rather, IFN-β impaired adhesion to, and spreading on, fibronectin, which was dependent on α5β1 (VLA-5)-integrin. IFN-β affected the function of VLA-5 in mature CACs, leading to rounding and detachment of cells, by induction of calpain 1 activity. Cell rounding and detachment was completely reversed by inhibition of calpain 1 activity in mature CACs. During in vitro capillary formation, CAC addition and calpain 1 inhibition enhanced sprouting of endothelial cells to a comparable extent, but were not sufficient to rescue tube formation in the presence of IFN-β. We show that the IFN-β-induced reduction of the numbers of in vitro differentiated CACs is based on activation of calpain 1, resulting in an attenuated adhesion to extracellular matrix proteins via VLA-5. In vivo, this could lead to inhibition of vessel formation due to reduction of the locally recruited CAC numbers and their paracrine angiogenic factors.
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http://dx.doi.org/10.1152/ajpheart.00810.2014DOI Listing
November 2015

Palmitic acid increases pro-oxidant adaptor protein p66Shc expression and affects vascularization factors in angiogenic mononuclear cells: Action of resveratrol.

Vascul Pharmacol 2015 Dec 5;75:7-18. Epub 2015 Aug 5.

Department of Molecular Cell Biology, VU University Medical Center, van der Boechorstraat 7, 1081BT Amsterdam, The Netherlands. Electronic address:

A defect in neo-vascularization process involving circulating angiogenic mononuclear cells (CACs) dysfunction is associated with diabetes. We showed that oxidative stress was elevated in CACs cultured from blood of individuals with metabolic syndrome (MetS) and diabetes. We then assessed the action of palmitic acid (PA), a deregulated and increased NEFA in metabolic disorders, focusing on its oxidant potential. We observed that the phyto-polyphenol resveratrol normalized oxidative stress both in CACs isolated from MetS patients or treated with PA. Resveratrol further decreased the deleterious action of PA on gene expression of vascularization factors (TNFα, VEGF-A, SDF1α, PECAM-1, VEGFR2, Tie2 and CXCR4) and improved CAC motility. Particularly, resveratrol abolished the PA-induced over-expression of the pro-oxidant protein p66Shc. Neither KLF2 nor SIRT1, previously shown in resveratrol and p66Shc action, was directly involved. Silencing p66Shc normalized PA action on VEGF-A and TNFα specifically, without abolishing the PA-induced oxidative stress, which suggests a deleterious role of p66Shc independently of any major modulation of the cellular oxidative status in a high NEFA levels context. Besides showing that resveratrol reverses PA-induced harmful effects on human CAC function, certainly through profound cellular modifications, we establish p66Shc as a major therapeutic target in metabolic disorders, independent from glycemic control.
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http://dx.doi.org/10.1016/j.vph.2015.08.003DOI Listing
December 2015

The diverse identity of angiogenic monocytes.

Eur J Clin Invest 2013 Jan 20;43(1):100-7. Epub 2012 Oct 20.

Department of Molecular Cell Biology and Immunology, VU University Medical Center, Amsterdam, the Netherlands.

Background: The role of bone marrow-derived cells in stimulating angiogenesis, vascular repair or remodelling has been well established, but the nature of the circulating angiogenic cells is still controversial.

Design: The existing literature on different cell types that contribute to angiogenesis in multiple pathologies, most notably ischaemic and tumour angiogenesis, is reviewed, with a focus on subtypes of angiogenic mononuclear cells and their local recruitment and activation.

Results: A large number of different cells of myeloid origin support angiogenesis without incorporating permanently into the newly formed vessel, which distinguishes these circulating angiogenic cells (CAC) from endothelial progenitor cells (EPC). Although CAC frequently express individual endothelial markers, they all share multiple characteristics of monocytes and only express a limited set of discriminative surface markers in the circulation. When cultured ex vivo, or surrounding the angiogenic vessel in vivo, however, many of them acquire similar additional markers, making their discrimination in situ difficult.

Conclusion: Different subsets of monocytes show angiogenic properties, but the distinct microenvironment, in vitro or in vivo, is needed for the development of their pro-angiogenic function.
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http://dx.doi.org/10.1111/eci.12009DOI Listing
January 2013

Coronary endothelial dysfunction after cardiomyocyte-specific mineralocorticoid receptor overexpression.

Am J Physiol Heart Circ Physiol 2011 Jun 25;300(6):H2035-43. Epub 2011 Mar 25.

Inserm U644, UFR Médecine-Pharmacie, 22 Boulevard Gambetta, 76183 Rouen Cedex, France.

The deleterious effects of aldosterone excess demonstrated in cardiovascular diseases might be linked in part to coronary vascular dysfunction. However, whether such vascular dysfunction is a cause or a consequence of the changes occurring in the cardiomyocytes is unclear. Moreover, the possible link between mineralocorticoid receptor (MR)-mediated effects on the cardiomyocyte and the coronary arteries is unknown. Thus we used a mouse model with conditional, cardiomyocyte-specific overexpression of human MR (hMR) and observed the effects on endothelial function in isolated coronary segments. hMR overexpression decreased the nitric oxide (NO)-mediated relaxing responses to acetylcholine in coronary arteries (but not in peripheral arteries), and this was prevented by a 1-mo treatment either with an MR antagonist, vitamin E/vitamin C, or a NADPH oxidase inhibitor. hMR overexpression did not affect coronary endothelial NO synthase content nor its level of phosphorylation on serine 1177, but increased cardiac levels of reactive oxygen species, cardiac NADPH oxidase (NOX) activity, and expression of the NOX subunit gp91phox, which was limited to endothelial cells. Thus an increase in hMR activation, restricted to cardiomyocytes, is sufficient to induce a severe coronary endothelial dysfunction. We suggest a new paracrine mechanism by which cardiomyocytes trigger a NOX-dependent, reactive oxygen species-mediated coronary endothelial dysfunction.
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http://dx.doi.org/10.1152/ajpheart.00552.2010DOI Listing
June 2011

Endothelial estrogen receptor {alpha} plays an essential role in the coronary and myocardial protective effects of estradiol in ischemia/reperfusion.

Arterioscler Thromb Vasc Biol 2010 Dec 16;30(12):2562-7. Epub 2010 Sep 16.

Department of Pharmacology, Institut National de la Santé et de la Recherche Médicale U644 and Rouen University Hospital, Institute for Biomedical Research Institut Fédératif de Recherches, University of Rouen, France.

Objective: To assess the coronary endothelial protective effects of 17β-estradiol (E2) and the role of estrogen receptor (ER) α in ischemia/reperfusion (I/R).

Methods And Results: E2 exerts protective effects in cardiac I/R. However, the implication in vivo of the endothelium and the cellular targets of the anti-ischemic effects of E2 are unknown. Mice were subjected to I/R (30 minutes of I and 1 hour of R) in vivo, after which acetylcholine-induced relaxation of isolated coronary segments was assessed ex vivo. I/R induced a coronary endothelial dysfunction in untreated ovariectomized mice that was prevented by long-term treatment with E2 in wild-type, but not in ERα(-/-), mice. Chimeric mice inactivated for ERα in the hematopoietic compartment remained protected by E2. Further inactivation of endothelial ERα abolished the protective action of E2 on coronary endothelial function in Tie2-Cre(+) ERα(f/f) mice. More importantly, E2 significantly limited infarct size in wild-type mice but not in mice deficient in endothelial ERα, even in the presence of hematopoietic ERα.

Conclusions: Endothelial ERα plays a crucial role in the E2-induced prevention of endothelial dysfunction after I/R. To our knowledge, we demonstrate for the first time, by using unique genetically modified mice, that targeting endothelial protection per se can confer cardiomyocyte protection in I/R.
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http://dx.doi.org/10.1161/ATVBAHA.110.213637DOI Listing
December 2010

Estrogen receptors and endothelium.

Arterioscler Thromb Vasc Biol 2010 Aug;30(8):1506-12

INSERM U858-I2MC, Faculté de Médecine, Université de Toulouse et Centre Hospitalier Universitaire, de Toulouse, BP 84225, 31432 Toulouse Cedex 4, France.

Estrogens, and in particular 17beta-estradiol (E2), play a pivotal role in sexual development and reproduction and are also implicated in a large number of physiological processes, including the cardiovascular system. Both acetylcholine-induced and flow-dependent vasodilation are preserved or potentiated by estrogen treatment in both animal models and humans. Indeed, E2 increases the endothelial production of nitric oxide and prostacyclin and prevents early atheroma through endothelial-mediated mechanisms. Furthermore, whereas it prevents endothelial activation, E2 potentiates the ability of several subpopulations of the circulating or resident immune cells to produce proinflammatory cytokines. The balance between these 2 actions could determine the final effect in a given pathophysiological process. E2 also promotes endothelial healing, as well as angiogenesis. Estrogen actions are essentially mediated by 2 molecular targets: estrogen receptor-alpha (ERalpha) and ERbeta. The analysis of mouse models targeted for ERalpha or ERbeta demonstrated a prominent role of ERalpha in vascular biology. ERalpha directly modulates transcription of target genes through 2 activation functions (AFs), AF-1 and AF-2. Interestingly, an AF-1-deficient ERalpha isoform can be physiologically expressed in the endothelium and appears sufficient to mediate most of the vasculoprotective actions of E2. In contrast, AF-1 is necessary for the E2 actions in reproductive targets. Thus, it appears conceivable to uncouple the vasculoprotective and sexual actions with appropriate selective ER modulators.
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http://dx.doi.org/10.1161/ATVBAHA.109.191221DOI Listing
August 2010

Reduced cardiac remodelling and prevention of glutathione deficiency after omega-3 supplementation in chronic heart failure.

Fundam Clin Pharmacol 2011 Jun;25(3):323-32

INSERM U644, Institut Fédératif de Recherches Multidisciplinaires sur les Peptides n°23, Rouen Institut for Biomedical Research and Inovation, UFR de Médecine et de Pharmacie, 22 Boulevard Gambetta, 76183 Rouen, France.

n-3 polyunsaturated fatty acids (omega-3) supplementation is associated with reduced cardiovascular mortality and post-infarction death. However, the impact of omega-3 supplementation in congestive heart failure (CHF) is still unknown. This study assesses the effects of omega-3 supplementation on left ventricular (LV) function and remodelling. We assessed, in rats with CHF induced by left coronary ligation, the effects of a 1-week and a 12-week supplementation with omega-3 (450 mg/kg per day) on LV hemodynamics, function and structure. Chronic omega-3 reduces total peripheral resistance due to an increase in cardiac output without modification of arterial pressure. Only chronic omega-3 reduces LV end-diastolic pressure and LV relaxation constant. Moreover, chronic omega-3 decreases LV systolic and diastolic diameters, LV weight and collagen density. Acute and chronic omega-3 increase LV γ-glutamyl-cysteine synthetase and oppose glutathione deficiency resulting in a reduction of myocardial oxidized glutathione. In experimental CHF, long-term omega-3 supplementation improves LV hemodynamics and function and prevents LV remodelling and glutathione deficiency. The latter might be one of the mechanisms involved, but whether other mechanism, independent of myocardial redox 'status', such as reduced inflammation, are implicated remains to be confirmed.
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http://dx.doi.org/10.1111/j.1472-8206.2010.00839.xDOI Listing
June 2011

Arterial stiffness is regulated by nitric oxide and endothelium-derived hyperpolarizing factor during changes in blood flow in humans.

Hypertension 2010 Mar 18;55(3):674-80. Epub 2010 Jan 18.

Department of Pharmacology, Rouen University Hospital and Institut National de la Sante et de la Recherche Medicale U644, Rouen Medical School, Institut Federatif de Recherche Multidisciplinaire sur les Peptides 23, Institute for Biomedical Research, University of Rouen, Rouen, France.

Cytochrome-derived epoxyeicosatrienoic acids may be important endothelium-derived hyperpolarizing factors, opening calcium-activated potassium channels, but their involvement in the regulation of arterial stiffness during changes in blood flow in humans is unknown. In healthy volunteers, we measured arterial pressure, radial artery diameter, wall thickness, and flow (NIUS02) during hand skin heating in the presence of saline or inhibitors of NO synthase (N(G)-monomethyl-L-arginine), calcium-activated potassium channels (tetraethylammonium), and cytochrome epoxygenases (fluconazole). Arterial compliance and elastic modulus were calculated and fitted as functions of midwall stress to suppress the confounding influence of geometric changes. Under saline infusion, heating induced an upward shift of the compliance-midwall stress curve and a downward shift of the modulus-midwall stress curve demonstrating a decrease in arterial tone and stiffness when blood flow increases. These shifts were reduced by N(G)-monomethyl-L-arginine and abolished by the combinations of N(G)-monomethyl-L-arginine+tetraethylammonium and N(G)-monomethyl arginine+fluconazole. In parallel, in isolated mice coronary arteries, fluconazole and tetraethylammonium reduced the relaxations to acetylcholine. However, fluconazole did not affect the relaxations to the openers of calcium-activated potassium channels of small- and intermediate-conductance NS309 and of large-conductance NS1619 excluding a direct effect on these channels. Moreover, tetraethylammonium reduced the relaxations to NS1619 but not to NS309, suggesting that the endothelium-derived hyperpolarizing factor involved mainly acts on large-conductance calcium-activated potassium channels. These results show in humans that, during flow variations, arterial stiffness is regulated by the endothelium through the release of both NO and cytochrome-related endothelium-derived hyperpolarizing factor.
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http://dx.doi.org/10.1161/HYPERTENSIONAHA.109.142190DOI Listing
March 2010

Toll-like receptors 4 contribute to endothelial injury and inflammation in hemorrhagic shock in mice.

Crit Care Med 2009 May;37(5):1724-8

Inserm U644, Institute for Biomedical Research and IFRMP 23, Rouen University Medical School, Rouen, France.

Objective: Hemorrhagic shock followed by resuscitation (HS/R) promotes organ injury by priming cells of the innate immune system for inflammatory response. Toll-like receptors (TLRs) play an important role in signal transduction in shock/resuscitation conditions. Because proinflammatory mediators are a critical event in mesenteric endothelial injury induced by HS/R, we assessed the role of TLR4 or TLR2 in this setting.

Design: Laboratory investigation.

Setting: Research laboratory at Rouen University Medical School.

Subjects: Male wild-type, TLR4(-/-) and TLR2(-/-) mice with the same C57BL/6 background.

Interventions: Mice were submitted to 30 minutes hemorrhagic shock followed by 1 hour resuscitation, after which mesenteric endothelial dysfunction, microvascular injury, and TNF[alpha] production were assessed.

Measurements And Main Results: HS/R markedly decreased nitric oxide-mediated mesenteric relaxations induced by acetylcholine, assessed ex vivo on a myograph. By contrast, in TLR4-deficient mice, HS/R did not impair the nitric oxide-mediated responses to acetylcholine. No protection was observed in TLR2-deficient mice. TLR4-deficient mice also displayed a significant reduction in fluid resuscitation and TNF[alpha] systemic production.

Conclusions: TLR4 contributes to mesenteric endothelial dysfunction after hemorrhagic shock. This early TLR4-induced vascular injury may be an important trigger of the systemic inflammatory response occurring in this disease.
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http://dx.doi.org/10.1097/CCM.0b013e31819da805DOI Listing
May 2009

Prevention of skin flap necrosis by estradiol involves reperfusion of a protected vascular network.

Circ Res 2009 Jan 4;104(2):245-54, 12p following 254. Epub 2008 Dec 4.

Institut National de la Santé et de la Recherche Médicale, U858, Toulouse, France.

Although 17beta-estradiol (E2) is protective in experimental models of myocardial and brain ischemia, its effect on skin ischemia remains unknown. Here, we assessed the protective effect of E2 in a mouse model of skin ischemia, mimicking the surgery of skin flaps. Whereas necrosis appeared in the half portion of the skin flap within 1 week after surgery in ovariectomized mice, it was reduced up to 10-fold when mice were pretreated with E2, at least 3 days before the surgery. The beneficial effect of E2 appeared to be attributable to an increase in skin survival, revealed by measuring viability of ex vivo explants and enhancement of the antiapoptotic Bcl-2 protein expression in vivo. This protective effect on the skin contributed to the protection of the vascular network and facilitated reperfusion, which was found to be accelerated in ovariectomized E2-treated mice, whereas hemorrhages were observed in untreated mice. E2 also increased expression of fibroblast growth factor-2 isoforms in the skin and circulating vascular endothelial growth factor in the serum. Finally, this protective effect of E2 was abolished in estrogen receptor-deficient mice (ERalpha(-/-)) but maintained in chimeric mice reconstituted with ERalpha-deficient bone marrow, indicating dispensable action of E2 in bone marrow-derived cells. This protective effect of E2 was mimicked by treatment with tamoxifen, a selective estrogen receptor modulator. In conclusion, we have demonstrated for the first time that E2 exerts a major preventive effect of skin flap necrosis through a prevention of ischemic-induced skin lesions, including those of the vascular network, which contributes to accelerate the reperfusion of the skin flap.
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http://dx.doi.org/10.1161/CIRCRESAHA.108.182410DOI Listing
January 2009

Role of endogenous endothelin in endothelial dysfunction in murine model of systemic sclerosis: tight skin mice 1.

Fundam Clin Pharmacol 2008 Dec;22(6):649-55

Inserm U644 & Rouen University Hospital, Institute for Biomedical Research and IFRMP 23, University of Rouen, Rouen, France.

Systemic sclerosis (SSc) is a systemic inflammatory disorder, resulting in severe vascular dysfunction. The endothelin (ET) system has vasoconstrictor and profibrotic properties and has been shown to be activated in SSc. ET antagonists are currently used in SSc-related pulmonary arterial hypertension, but the endothelial impact of ET antagonists remains less known in SSc. We thus assessed the effects of the dual ET(A)-ET(B) antagonist, bosentan, on endothelial dysfunction in a murine model of SSc, the heterozygous tight-skin mice 1 (TSK1(+)). Six-week-old TSK1(+) were either untreated or treated for 6 weeks with bosentan (100 mg/kg/day), and compared with controls. Endothelial function was evaluated in isolated mesenteric resistance arteries, using a small vessel myograph. TSK1(+) displayed endothelial dysfunction, as shown by a decreased response of mesenteric arteries to acetylcholine, especially in the presence of L-nitro-arginine methyl ester (L-NAME), corresponding to NO-independent, prostaglandin-mediated relaxation. The NO-independent relaxation was partially restored in bosentan-treated TSK1(+), and this was abolished by a cyclo-oxygenase inhibitor. Therefore, the murine model of SSc, TSK1(+) exhibits severe endothelial dysfunction of peripheral resistance arteries. The ET antagonist bosentan prevents endothelial alterations, suggesting a major role of ET in the adverse vascular effects of SSc.
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http://dx.doi.org/10.1111/j.1472-8206.2008.00634.xDOI Listing
December 2008

Aldosterone synthase inhibition improves cardiovascular function and structure in rats with heart failure: a comparison with spironolactone.

Eur Heart J 2008 Sep 27;29(17):2171-9. Epub 2008 Jun 27.

Faculté de Médecine et Pharmacie, INSERM U644, 22 Boulevard Gambetta, 76183 Rouen Cedex, France.

Aims: Inhibition of aldosterone synthase, the key enzyme in aldosterone formation, could be an alternative strategy for mineralocorticoid-receptor antagonists in congestive heart failure (CHF), but its effect in CHF is unknown.

Methods And Results: We compared, in rats with CHF, the effects of a 7 day and a 12 week treatment with the aldosterone synthase inhibitor FAD286 (4 mg kg(-1) day(-1)) with those induced by spironolactone (80 mg kg(-1) day(-1)). FAD286/spironolactone increased cardiac output without modifying arterial pressure. Long-term FAD286 and spironolactone reduced left ventricular (LV) end-diastolic pressure, LV relaxation constant, and LV dilatation, and these effects were more marked with FAD286, whereas both drugs reduced LV hypertrophy and collagen accumulation to the same extent. Long-term FAD286/spironolactone prevented CHF-related enhancement in LV ACE and reduction in LV ACE-2, but only FAD286 prevented the reduction in LV AT(2) receptors. FAD286, but not long-term spironolactone, reduced the CHF-related enhancements in LV reactive oxygen species, reduced-oxidized glutathione ratio, and aortic nicotinamide adenine dinucleotide phosphate oxidase activity. FAD286 normalized the CHF-induced impairment of endothelium-dependent vasodilatation.

Conclusion: In experimental CHF, FAD286 and spironolactone improve LV haemodynamics, remodelling, and function, but only FAD286 persistently normalizes LV 'redox status'. These results suggest that aldosterone synthase inhibition is a potential therapeutic strategy for the treatment of CHF.
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http://dx.doi.org/10.1093/eurheartj/ehn277DOI Listing
September 2008

Aldosterone-induced coronary dysfunction in transgenic mice involves the calcium-activated potassium (BKCa) channels of vascular smooth muscle cells.

Circulation 2007 Nov 5;116(21):2435-43. Epub 2007 Nov 5.

Inserm U689 and Paris Diderot University, Paris, France.

Background: Cardiomyocyte-specific overexpression of aldosterone synthase in male (MAS) mice induces a nitric oxide-independent coronary dysfunction. Because calcium-activated potassium (BKCa) channels are essential for vascular smooth muscle cell (VSMC) relaxation, we hypothesized that aldosterone alters their expression and/or function in VSMCs.

Methods And Results: Left coronary artery segments were isolated from MAS or male wild-type mice and mounted in a wire myograph. Responses to acetylcholine were assessed (in the presence of a nitric oxide synthase inhibitor) without or with the cyclooxygenase inhibitor diclofenac, the KCa inhibitors charybdotoxin plus apamin, or the BKCa inhibitor iberiotoxin. Expression of BKCa was quantified in hearts by real-time quantitative polymerase chain reaction and Western blot and in isolated coronary arteries by polymerase chain reaction. The effect of aldosterone on BKCa expression also was studied in cultured rat aortic VSMCs. Acetylcholine-mediated coronary relaxation was markedly decreased in MAS mice and was prevented by spironolactone. Diclofenac did not affect the MAS-induced impairment in the responses to acetylcholine, whereas charybdotoxin plus apamin virtually abolished the relaxation in both male wild-type and MAS mice. After iberiotoxin, relaxation to acetylcholine was decreased to a larger extent in male wild-type than in MAS, leading to similar levels of relaxation. BKCa-alpha and -beta1 subunit expressions were significantly decreased in MAS heart and coronary arteries. In cultured VSMCs, aldosterone induced a concentration-dependent decrease in BKCa expression, which was prevented by spironolactone.

Conclusions: Aldosterone overexpression altered VSMC BKCa expression and coronary BKCa-dependent relaxation. The resulting alteration of relaxing responses may contribute to the deleterious effects of aldosterone in cardiovascular diseases. BKCa channels may therefore be useful therapeutic targets in cardiovascular diseases.
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http://dx.doi.org/10.1161/CIRCULATIONAHA.107.722009DOI Listing
November 2007

Sonic hedgehog carried by microparticles corrects endothelial injury through nitric oxide release.

FASEB J 2007 Sep 11;21(11):2735-41. Epub 2007 Apr 11.

INSERM, U771, CNRS, UMR 6214, Université d'Angers, Faculté de Médecine, Rue Haute de Reculée, Angers, F-49045 France.

Microparticles (MPs) are small fragments generated from the plasma membrane after cell stimulation. Among the candidate proteins harbored by MPs, we recently showed that sonic hedgehog (Shh) is present in MPs generated from activated/apoptotic human T lymphocytes [Martínez et al., Blood (2006) vol. 108, 3012-3020]. We show here that Shh carried by MPs induces nitric oxide (NO) release from endothelial cells, triggers changes in the expression and phosphorylation of enzymes related to the NO pathway, and decreases production of reactive oxygen species. When PI3-kinase and ERK signaling were specifically inhibited, the effects of MPs were reversed. In vivo injection of MPs in mice was also able to improve endothelial function by increasing NO release, and it reversed endothelial dysfunction after ischemia/reperfusion. Silencing the effects of Shh with cyclopamine, a specific inhibitor of Shh, or siRNA, an inhibitor of the Shh receptor Patched, strongly reduced production of NO elicited by MPs. Taken together, we propose that the biological message carried by MPs harboring Shh may represent a new therapeutic approach against endothelial dysfunction during acute severe endothelial injury.
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http://dx.doi.org/10.1096/fj.07-8079comDOI Listing
September 2007

Toll-like receptors 2-deficient mice are protected against postischemic coronary endothelial dysfunction.

Arterioscler Thromb Vasc Biol 2007 May 1;27(5):1064-71. Epub 2007 Mar 1.

Inserm U644 & Rouen University Hospital, Institute for Biomedical Research and IFRMP 23, University of Rouen, France.

Objectives: Toll-like receptors (TLR) 2 are expressed in cardiac and inflammatory cells, and regulate leukocyte function. Because leukocyte adhesion is a critical event in endothelial injury induced by ischemia/reperfusion (I/R), we assessed whether TLR2 were involved in I/R-induced coronary endothelial injury.

Methods And Results: Ischemia-reperfusion markedly decreased NO-mediated coronary relaxations to acetylcholine assessed ex vivo. In contrast, in TLR2 deficient mice, I/R paradoxically improved the NO-mediated responses to acetylcholine. To precise the cellular compartment expressing TLR2 which is involved in endothelial injury, we developed bone-marrow chimeric mice by transplanting TLR2-/- bone marrow to WT mice or WT bone marrow to TLR2-/- mice and submitted them to I/R 5 weeks after transplant. Both chimeric mice displayed similar protection as TLR2-/- mice against I/R-induced endothelial dysfunction, suggesting a role of TLR2 expressed on both non-bone marrow cells (in our case presumably endothelial cells and/or cardiomyocytes) and cells of bone marrow origin (presumably neutrophils). TLR2 deficiency was also associated with a smaller infarct size, and reduced reperfusion-induced production of reactive oxygen species and leukocyte infiltration.

Conclusions: TLR2 contribute to coronary endothelial dysfunction after I/R, possibly through stimulation of neutrophil- (and free radical-) mediated endothelial injury.
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http://dx.doi.org/10.1161/ATVBAHA.107.140723DOI Listing
May 2007

NO produced by endothelial NO synthase is a mediator of delayed preconditioning-induced endothelial protection.

Am J Physiol Heart Circ Physiol 2003 Jun 9;284(6):H2053-60. Epub 2003 Jan 9.

Institut National de la Santé et de la Recherche Médicale E9920, Department of Pharmacology, Rouen University Medical School, France.

Preconditioning with brief periods of ischemia-reperfusion (I/R) induces a delayed protection of coronary endothelial cells against reperfusion injury. We assessed the possible role of nitric oxide (NO) produced during prolonged I/R as a mediator of this endothelial protection. Anesthetized rats were subjected to 20-min cardiac ischemia/60-min reperfusion, 24 h after sham surgery or cardiac preconditioning (1 x 2-min ischemia/5-min reperfusion and 2 x 5-min ischemia/5-min reperfusion). The nonselective NO synthase (NOS) inhibitor l-NAME, the selective inhibitors of neuronal (7-nitroindazole) or inducible (1400W) NOS, or the peroxynitrite scavenger seleno-l-methionine were administered 10 min before prolonged ischemia. Preconditioning prevented the reperfusion-induced impairment of coronary endothelium-dependent relaxations to acetylcholine (maximal relaxation: sham 77 +/- 3; I/R 44 +/- 6; PC 74 +/- 5%). This protective effect was abolished by l-NAME (41 +/- 7%), whereas 7-NI, 1400W or seleno-l-methionine had no effect. The abolition of preconditioning by l-NAME, but not by selective nNOS or iNOS inhibition, suggests that NO produced by eNOS is a mediator of delayed endothelial preconditioning.
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http://dx.doi.org/10.1152/ajpheart.00627.2002DOI Listing
June 2003