Temporal cross talk between endoplasmic reticulum and mitochondria regulates oxidative stress and mediates microparticle-induced endothelial dysfunction

Aims: Circulating microparticles (MPs) from metabolic syndrome patients and those generated from apoptotic T-cells induce endothelial dysfunction; however, the molecular and cellular mechanism(s) underlying in the effects of MPs remain to be elucidated. Results: Here, we show that both types of MPs increased expression of endoplasmic reticulum (ER) stress markers XBP-1, p-eIF2alpha and CHOP and nuclear translocation of ATF6 on human aortic endothelial cells. MPs decreased in vitro nitric oxide release by human aortic endothelial cells, whereas in vivo MP injection into mice impaired the endothelium-dependent relaxation induced by acetylcholine. These effects were prevented when ER stress was inhibited suggesting that ER stress is implicated in the endothelial effects induced by MPs. MPs affected mitochondrial function and evoked sequential increase of cytosolic and mitochondrial reactive oxygen species (ROS). Pharmacological inhibition of ER stress and silencing of neutral sphingomyelinase with siRNA abrogated all MP-mediated effects. Neutralization of Fas-Ligand carried by MPs abolished effects induced by both MP types, whereas neutralization of low density lipoprotein-receptor on endothelial cells prevented T-lymphocyte MP-mediated effects. Innovation and Conclusion: Collectively, endothelial dysfunction triggered by MPs involves temporal cross-talk between ER and mitochondria with respect to spatial regulation of ROS via the neutral sphingomyelinase and interaction of MPs with Fas and/or low density lipoprotein-receptor. These results provide a novel molecular insight into the manner MPs mediate vascular dysfunction and allow identification of potential therapeutic targets to treat vascular complications associated with metabolic syndrome.Institut National de la Santé et de la Recherche Médicale, Université dʼAngers and Centre Hospitalo-Universitaire dʼAngers

Microparticles Carrying Sonic Hedgehog Favor Neovascularization through the Activation of Nitric Oxide Pathway in Mice

BACKGROUND: Microparticles (MPs) are vesicles released from plasma membrane upon cell activation and during apoptosis. Human T lymphocytes undergoing activation and apoptosis generate MPs bearing morphogen Shh (MPs(Shh+)) that are able to regulate in vitro angiogenesis.METHODOLOGY/PRINCIPAL FINDINGS: Here, we investigated the ability of MPs(Shh+) to modulate neovascularization in a model of mouse hind limb ischemia. Mice were treated in vivo for 21 days with vehicle, MPs(Shh+), MPs(Shh+) plus cyclopamine or cyclopamine alone, an inhibitor of Shh signalling. Laser doppler analysis revealed that the recovery of the blood flow was 1.4 fold higher in MPs(Shh+)-treated mice than in controls, and this was associated with an activation of Shh pathway in muscles and an increase in NO production in both aorta and muscles. MPs(Shh+)-mediated effects on flow recovery and NO production were completely prevented when Shh signalling was inhibited by cyclopamine. In aorta, MPs(Shh+) increased activation of eNOS/Akt pathway, and VEGF expression, being inhibited by cyclopamine. By contrast, in muscles, MPs(Shh+) enhanced eNOS expression and phosphorylation and decreased caveolin-1 expression, but cyclopamine prevented only the effects of MPs(Shh+) on eNOS pathway. Quantitative RT-PCR revealed that MPs(Shh+) treatment increased FGF5, FGF2, VEGF A and C mRNA levels and decreased those of α5-integrin, FLT-4, HGF, IGF-1, KDR, MCP-1, MT1-MMP, MMP-2, TGFβ1, TGFβ2, TSP-1 and VCAM-1, in ischemic muscles. CONCLUSIONS/SIGNIFICANCE: These findings suggest that MPs(Shh+) may contribute to reparative neovascularization after ischemic injury by regulating NO pathway and genes involved in angiogenesis

Measurement of proton, deuteron, triton, and α particle emission after nuclear muon capture on Al, Si, and Ti with the AlCap experiment

Heavy charged particles after nuclear muon capture are an important nuclear physics background to the muon-to-electron conversion experiments Mu2e and COMET, which will search for charged lepton flavor violation at an unprecedented level of sensitivity. The AlCap experiment measured the yield and energy spectra of protons, deuterons, tritons, and alpha particles emitted after the nuclear capture of muons stopped in Al, Si, and Ti in the low energy range relevant for the muon-to-electron conversion experiments. Individual charged particle types were identified in layered silicon detector packages and their initial energy distributions were unfolded from the observed energy spectra. Detailed information on yields and energy spectra for all observed nuclei are presented in the paper.Comment: 24 pages, 19 figure

Rôle des microparticules membranaires dans l'angiogenèse

Microparticles (MPs) are small vesicles released from the plasma membrane after cell stimulation or apoptosis. Recent studies show that MPs can be implicated in modulation of angiogenesis, an essential cellular process from cardiovascular system during physiological and pathological conditions. Moreover, engineered MPs generated from apoptotic/stimulated human lymphocytes harbour at their surface the morphogen Sonic Hedgehog (Shh) which also play a critical role in vessel formation. Accordingly, the present study was aimed to examine how engineered human MPs bearing Shh (MPsShh+) are able to modulate the different steps implicated in angiogenesis process. In human endothelial cells (ECs), MPsShh+ induce the formation of capillary-like structures. Moreover, MPsShh+ are able to modulate EC proliferation, adhesion and mRNA levels of proangiogenic factors. Silencing of Shh receptor, by siRNA or Shh signalling by pharmacological inhibitor cyclopamine, strongly reverses tubulogenesis, and concomitantly decreases mRNA levels of pro-angiogenic factors. Furthermore, we investigate the effects of MPsShh+ on post-ischemic neovascularization using mice model of femoral artery ligation. The recovery of ischemic/normal blood flow ratio is greater in MPsShh+-treated mice than in control. In muscle, MPsShh+ enhance the expression of eNOS, decrease expression of caveolin-1 and modify expression of several angiogenic genes. Pharmacological inhibition of Shh pathway completely reverses the improvement of blood flow and the effects on eNOS expression. Besides, in light of our results, we proposed the hypothesis that MPsShh+ may represent a new therapeutic approach to treat pathologies associated with failed angiogenesis and a new target in pathologies associated with exaggeration of angiogenesis, such as cancer.Les microparticules (MPs) sont des fragments de la membrane plasmique libérés suite à une stimulation cellulaire ou apoptotique. Des études récentes ont montré que les MPs peuvent être impliquées dans la modulation de l'angiogenèse, un processus essentiel du système cardio-vasculaire pendant les états physiologiques et pathologiques. De plus, les MPs dérivées des cellules T humaines activées et apoptotiques sont porteuses du morphogène Sonic Hedgehog (Shh), lequel a aussi un rôle dans la formation des vaisseaux. En conséquence, l'objectif de cette étude est d'élucider l'implication des MPs porteuses de Shh (MPsShh+) sur les différentes étapes associées à l'angiogenèse. Dans les cellules endothéliales, les MPsShh+ provoquent la formation des structures capillaires. Également, les MPsShh+ sont capables de moduler la prolifération, l'adhésion et le niveau d'expression des ARNm des facteurs pro-angiogéniques dans les cellules endothéliales. De plus, l'inhibition de l'expression du récepteur de Shh, ou l'utilisation de la cyclopamine, un inhibiteur pharmacologique de Shh, restitue la capacité des MPsShh+ de former des capillaires et aussi diminue l'expression des facteurs pro-angiogéniques. Ensuite, nous avons évalué les effets des MPsShh+ sur la neovascularization post-ischémique en utilisant le modèle in vivo de la ligature de l'artère fémorale chez la souris. Une meilleure récupération du flux sanguin est observée chez les souris traitées avec les MPsShh+ par rapport aux témoins. Dans le muscle squelettique, les MPsShh+ augmentent l'expression de la eNOS et diminuent celle de la caveoline-1. L'inhibition pharmacologique de la voie Shh prévient l'amélioration du flux sanguin et les effets induits sur l'expression de la eNOS. L'ensemble des résultats suggère que les MPsShh+ pourraient être utilisées comme de nouveaux outils thérapeutiques dans certaines pathologies caractérisées par une angiogenèse défaillante et elles pourraient représenter une nouvelle cible dans les pathologies associées à une angiogenèse exagérée, comme le cancer

Rôle des microparticules membranaires dans l'angiogenèse

Les microparticules (MPs) sont des fragments de la membrane plasmique libérés suite à une stimulation cellulaire ou apoptotique. Des études récentes ont montré que les MPs peuvent être impliquées dans la modulation de l'angiogenèse, un processus essentiel du système cardio-vasculaire pendant les états physiologiques et pathologiques. De plus, les MPs dérivées des cellules T humaines activées et apoptotiques sont porteuses du morphogène Sonic Hedgehog (Shh), lequel a aussi un rôle dans la formation des vaisseaux. En conséquence, l'objectif de cette étude est d'élucider l'implication des MPs porteuses de Shh (MPsShh+) sur les différentes étapes associées à l'angiogenèse. Dans les cellules endothéliales, les MPsShh+ provoquent la formation des structures capillaires. Également, les MPsShh+ sont capables de moduler la prolifération, l'adhésion et le niveau d'expression des ARNm des facteurs pro-angiogéniques dans les cellules endothéliales. De plus, l'inhibition de l'expression du récepteur de Shh, ou l'utilisation de la cyclopamine, un inhibiteur pharmacologique de Shh, restitue la capacité des MPsShh+ de former des capillaires et aussi diminue l'expression des facteurs pro-angiogéniques. Ensuite, nous avons évalué les effets des MPsShh+ sur la neovascularization post-ischémique en utilisant le modèle in vivo de la ligature de l'artère fémorale chez la souris. Une meilleure récupération du flux sanguin est observée chez les souris traitées avec les MPsShh+ par rapport aux témoins. Dans le muscle squelettique, les MPsShh+ augmentent l'expression de la eNOS et diminuent celle de la caveoline-1. L'inhibition pharmacologique de la voie Shh prévient l'amélioration du flux sanguin et les effets induits sur l'expression de la eNOS. L'ensemble des résultats suggère que les MPsShh+ pourraient être utilisées comme de nouveaux outils thérapeutiques dans certaines pathologies caractérisées par une angiogenèse défaillante et elles pourraient représenter une nouvelle cible dans les pathologies associées à une angiogenèse exagérée, comme le cancer.Microparticles (MPs) are small vesicles released from the plasma membrane after cell stimulation or apoptosis. Recent studies show that MPs can be implicated in modulation of angiogenesis, an essential cellular process from cardiovascular system during physiological and pathological conditions. Moreover, engineered MPs generated from apoptotic/stimulated human lymphocytes harbour at their surface the morphogen Sonic Hedgehog (Shh) which also play a critical role in vessel formation. Accordingly, the present study was aimed to examine how engineered human MPs bearing Shh (MPsShh+) are able to modulate the different steps implicated in angiogenesis process. In human endothelial cells (ECs), MPsShh+ induce the formation of capillary-like structures. Moreover, MPsShh+ are able to modulate EC proliferation, adhesion and mRNA levels of proangiogenic factors. Silencing of Shh receptor, by siRNA or Shh signalling by pharmacological inhibitor cyclopamine, strongly reverses tubulogenesis, and concomitantly decreases mRNA levels of pro-angiogenic factors. Furthermore, we investigate the effects of MPsShh+ on post-ischemic neovascularization using mice model of femoral artery ligation. The recovery of ischemic/normal blood flow ratio is greater in MPsShh+-treated mice than in control. In muscle, MPsShh+ enhance the expression of eNOS, decrease expression of caveolin-1 and modify expression of several angiogenic genes. Pharmacological inhibition of Shh pathway completely reverses the improvement of blood flow and the effects on eNOS expression. Besides, in light of our results, we proposed the hypothesis that MPsShh+ may represent a new therapeutic approach to treat pathologies associated with failed angiogenesis and a new target in pathologies associated with exaggeration of angiogenesis, such as cancer.ANGERS-BU Médecine-Pharmacie (490072105) / SudocSudocFranceF

The Potential Neuroprotective Role of Free and Encapsulated Quercetin Mediated by miRNA against Neurological Diseases

International audienceChronic neuroinflammation is a pathological condition of numerous central nervous system (CNS) diseases such as Parkinson’s disease, Alzheimer’s disease, multiple sclerosis, amyotrophic lateral sclerosis and many others. Neuroinflammation is characterized by the microglia activation and concomitant production of pro-inflammatory cytokines leading to an increasing neuronal cell death. The decreased neuroinflammation could be obtained by using natural compounds, including flavonoids known to modulate the inflammatory responses. Among flavonoids, quercetin possess multiple pharmacological applications including anti-inflammatory, antitumoral, antiapoptotic and anti-thrombotic activities, widely demonstrated in both in vitro and in vivo studies. In this review, we describe the recent findings about the neuroprotective action of quercetin by acting with different mechanisms on the microglial cells of CNS. The ability of quercetin to influence microRNA expression represents an interesting skill in the regulation of inflammation, differentiation, proliferation, apoptosis and immune responses. Moreover, in order to enhance quercetin bioavailability and capacity to target the brain, we discuss an innovative drug delivery system. In summary, this review highlighted an important application of quercetin in the modulation of neuroinflammation and prevention of neurological disorders

Lymphocytic microparticles confer endothelial protection through activation of antioxidant defenses

National audienceOxidative stress plays an important role in the initial occurrence of cardiovascular diseases. Previous studies demonstrated that microparticles (MPs) released from activated and apoptotic lymphocytes improve endothelial function due to their ability to decrease reactive oxygen species (ROS). Here, we evaluated the capacity of these MPs to modulate ROS production during actinomicyn D (ActD)-induced apoptosis in human umbilical vein endothelial cells (HUVECs).MPs were generated from T lymphocytes undergoing activation with phytohemagglutinin and then, by stimulation with phorbol-12 myristate-13 acetate and ActD. HUVECs were grown for 24h in absence or presence of ActD (1g/ml), and/or 10Pg protein/ml of MPs.Once evaluated that MPs are able to prevent ActD-evoked apoptosis, we investigated the time course of modifcations in ROS production (i.e. 1, 2, 4, 6, 8, 10 and 24h) by electron paramagnetic resonance. ActD enhances ROS production after 2h and 10h of treatment that was prevented by MPs after 2h but not at 10h, indicating that they act during the early phase of apoptosis. Interestingly at 24h, superoxide dismutase (SOD) mimetic, manganese (III) tetrakis (1-methyl-4-pyridyl) porphyrin (MnTMPyP) decreases cell death and inhibition of apoptosis induced by MPs, suggesting that MPs may behave as an SOD mimetic. Although these MPs carry functionally active extracellular SOD and catalase, as demonstrated by western blot analysis, treatment of HUVECs with these MPs does not modify expression of these enzymes.Under the same experimental conditions, MPs increase the expressions of other antioxidant enzymes such as Cu/Zn SOD and Mn SOD in HUVECs that may explain the protective effects induced by MPs. Thus, the present study illustrates new mechanisms by which MPs from activated/apoptotic lymphocytes may confer endothelial protection under pathological conditions in which oxidative stress is enhanced by a subtle modulation of antioxidant defense mechanism.</p

Microparticles from T lymphocytes reduce actinomycin D-induced apoptosis in human endothelial cells

Date du colloque&nbsp;: 2009National audienc

Engineered microparticles bearing the morphogen Sonic Hedgehog protect endothelial cells against actinomycin D-induced apoptosis

Date du colloque&nbsp;: 04/2010National audienc