Microparticles are vectors of paradoxical information in vascular cells including the endothelium: role in health and diseases

Both inflammation and thrombosis can be orchestrated by the interactions between circulating cells, such as leukocytes and platelets, with vascular, endothelial and smooth muscle cells, which, during activation or apoptosis, can release circulating microparticles (MPs). Indeed, MPs are membrane vesicles with procoagulant and proinflammatory properties. MPs are present in blood from healthy individuals and in patients under several pathological states, for instance sepsis, preeclampsia, Crohn\u27s disease and diabetes, strengthening the notion that MPs may play a role in these diseases. Circulating MPs or those generated in vitro from apoptotic T cells display deleterious effects on endothelial and/or vasomotor function. In contrast, MPs might be protective to endothelial cells. We have shown that MPs harboring the morphogen sonic hedgehog may represent a new therapeutic approach against endothelial dysfunction during acute severe endothelial injury. Indeed, these types of MPs induce NO release, decrease production of reactive oxygen species and induce angiogenesis from endothelial cells. This protective role for the endothelium was confirmed also by their in vivo injection in mice in which they were also able to reverse endothelial dysfunction in a model of heart ischemia/reperfusion. On the contrary, MPs from preeclamptic women compared to those from normal pregnant women showed pro-inflammatory properties in the vascular wall inducing vascular hyporeactivity in vessels from humans and mice. These effects were associated with complex interactions between NO and cyclooxygenase systems via endothelial cell activation. Altogether, these findings suggest that MPs can be considered as vectors of biological messages for vascular homeostasis, during immunity and inflammation

Deletion of estrogen receptor-alpha abolishes endothelial response to wine polyphenols without affecting the main cardiovascular parameters in mice

Date du colloque : 10/2009International audienc

Cyclooxygenase-2-Derived Prostacyclin Protective Role on Endotoxin-Induced Mouse Cardiomyocyte Mortality

Cardiovascular dysfunction characterizes septic shock, inducing multiple organ failure and a high mortality rate. In the heart, it has been shown an up-regulation of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expressions with subsequent overproduction of nitric oxide (NO) and eicosanoids. This study is focused on the links between these products of inflammation and cell loss of mouse cardiomyocytes during treatment by the Salmonella typhimurium lipopolysaccharide (LPS) in presence or in absence of NOS or COX inhibitors. LPS induced RelA/NF-κB p65 activation, iNOS and COX-2 up-regulations, resulting in NO and prostacyclin releases. These effects were reversed by the NO-synthase inhibitor and increased by the specific COX-2 inhibitor. Immunostainings with FITC-conjugated anti-Annexin-V and propidium iodide and caspase 3/7 activity assay showed that cardiomyocyte necrosis was inhibited by L-NA during LPS treatment challenge, while apoptosis was induced in presence of both LPS and NS-398. No effect on LPS cellular injury was observed using the specific cyclooxygenase-1 (COX-1) inhibitor, SC-560. These findings strongly support the hypothesis of a link between iNOS-dependent NO overproduction and LPS-induced cell loss with a selective protective role allotted to COX-2 and deriving prostacyclins

Increase in circulating microparticles in inflammatory bowel disease patients induces vascular alterations

Alterations of vascular function participate to the development of the inflammatory bowel disease (IBD). We have previously reported the presence of inflammation-related vasoactive factors in small mesenteric arteries from Crohn’s disease patients (Tabernero et al., Circulation, 2003). Microparticles (MPs) are membrane vesicles released during cell activation and apoptosis whose circulating levels have been shown to be increased in patients with IBD, including MPs from platelets and activated platelets, endothelial, leukocyte and erythrocyte origins (Leonetti et al., Archives of Cardiovascular Diseases, Tome 102-Supplement 1-Mars 2009.).Here, we investigated whether MPs from IBD patients (IBD-MPs) play a role in the regulation of endothelial function and vascular reactivity in this disease. Male Swiss mice were injected intravenously with IBD-MPs or saline solution and sacrificed 24 h after. Endothelial function and vascular reactivity were studied on aortic rings by myography. The involvement of nitric oxide (NO), cyclooxygenase (COX) metabolites and superoxide anion (O2- ) was also assessed using the following inhibitors: NG-L-Nitro-arginine (NO synthase inhibitor, L-NA); indomethacin (non-selective COX inhibitor); SC-560 (selective COX-1 inhibitor), NS-398 (selective COX-2 inhibitor) and MnTMPyP (permeable superoxide dismutase mimetic). In aorta, IBD-MPs significantly reduced both endothelium-dependent induced by acetylcholine and the NO donor, sodium nitroprusside, respectively. IBD-MPs decreased the contraction to serotonin (5-HT) compared to saline that was completely prevented in the presence of L-NA. Moreover, aorta from mice treated with IBD-MPs displayed increase NO production. Interestingly, the ability of NS-398 to reduce 5-HT-induced contraction in control mice was abolished in vessels taken from mice treated with IBD-MPs. Although IBD-MPs decreased O2- production in the aorta, the O2- scavenger MnTMPyP reduced the contraction to 5-HT in an identical manner in aorta from both control and IBD-MPs treated mice. The present study provides evidence of the capacity of IBD-MPs to induce endothelial dysfunction and vascular hyporeactivity. These effects result from a subtle alteration of the balance between NO, reactive oxygen species and metabolites from COX-2. They underscore the participation of MPs in the course of vascular alterations in this disease. (Partially supported by Ferring France Laboratories)

Involvement of β3-Adrenoceptor in Altered β-Adrenergic Response in Senescent Heart: Role of Nitric Oxide Synthase 1–derived Nitric Oxide

Background: In senescent heart, β-adrenergic response is altered in parallel with β1- and β2-adrenoceptor down-regulation. A negative inotropic effect of β3-adrenoceptor could be involved. In this study, the authors tested the hypothesis that β3-adrenoceptor plays a role in β-adrenergic dysfunction in senescent heart.Methods: β-Adrenergic responses were investigated in vivo (echocardiography–dobutamine, electron paramagnetic resonance) and in vitro (isolated left ventricular papillary muscle, electron paramagnetic resonance) in young adult (3-month-old) and senescent (24-month-old) rats. Nitric oxide synthase (NOS) immunolabeling (confocal microscopy), nitric oxide production (electron paramagnetic resonance) and β-adrenoceptor Western blots were performed in vitro. Data are mean percentages of baseline ± SD. Results: An impaired positive inotropic effect (isoproterenol) was confirmed in senescent hearts in vivo (117 ± 23 vs. 162 ± 16%; P < 0.05) and in vitro (127 ± 10 vs. 179 ± 15%; P < 0.05). In the young adult group, the positive inotropic effect was not significantly modified by the nonselective NOS inhibitor NG-nitro-l-arginine methylester (l-NAME; 183 ± 19%), the selective NOS1 inhibitor vinyl-l-N-5(1-imino-3-butenyl)-l-ornithine (l-VNIO; 172 ± 13%), or the selective NOS2 inhibitor 1400W (183 ± 19%). In the senescent group, in parallel with β3-adrenoceptor up-regulation and increased nitric oxide production, the positive inotropic effect was partially restored by l-NAME (151 ± 8%; P < 0.05) and l-VNIO (149 ± 7%; P < 0.05) but not by 1400W (132 ± 11%; not significant). The positive inotropic effect induced by dibutyryl-cyclic adenosine monophosphate was decreased in the senescent group with the specific β3-adrenoceptor agonist BRL 37344 (167 ± 10 vs. 142 ± 10%; P < 0.05). NOS1 and NOS2 were significantly up-regulated in the senescent rat. Conclusions: In senescent cardiomyopathy, β3-adrenoceptor overexpression plays an important role in the altered β-adrenergic response via induction of NOS1-nitric oxide

Influence of micropaticles harvested from patients affected by obstructive sleep apnea syndrome on endothelial function and vascular reactivity

Obstructive sleep apnea syndrome (OSAS) is a highly prevalent disease characterized by recurrent episodes of partial or complete obstruction of the upper airways during sleep, leading to oxygen desaturation, sleep fragmentation and clinical endothelial dysfunction. Microparticles (MPs) are membrane vesicles released during cell activation and apoptosis. Elevated levels of circulating MPs have been detected in pathologies associated with vascular alterations. We investigated the effects of MPs on endothelial function and vascular reactivity in OSAS. Blood samples were obtained either from 38 OSAS patients without any other cardiovascular comorbidities and 23 healthy subjects. A desaturation index above 10 per hour defined OSAS patients. MPs concentration and origin were assessed using flow cytometer. Male Swiss mice were injected i.v. with MPs from OSAS or healthy subjects, or with saline solution, and sacrified after 24hours. Endothelial function and vascular reactivity were studied on aortic rings and small mesenteric resistance (SMA) arteries by myography and arteriography, respectively. Patients with OSAS did not display increased circulating levels of MPs compared to healthy subjects including those from pro-coagulant, platelet, endothelial, leukocyte and erythrocyte origins. Interestingly, MPs from granulocytes and activated leukocytes were significantly enhanced in OSAS patients. Activated leukocyte MPs positively correlated with oxygen desaturation index. In aorta, MPs from OSAS patients but not those from healthy subjects significantly reduced endothelium-dependent relaxation to acetylcholine. MPs from OSAS increased sensitivity of the aorta in response to serotonin that was greater compared to the effect of MPs from healthy subjects. In SMA, MPs from OSAS but not those from healthy subjects impaired flow-induced dilation without any effect on myogenic tone. Although SMA from mice treated with healthy subjects MPs did not affect flow-induced dilation, these vessels showed a reduced prostacyclin-component that was completely compensated by the NO-component of the response. The endothelial dysfunction induced by MPs from OSAS was caused by the reduction of both NO- and prostacyclin- but not the endothelium-derived hyperpolarizing factor-components of the response in SMA. These data provide evidence that circulating MPs from OSAS patients influence both endothelial function and vascular reactivity

Rosiglitazone, a Peroxisome Proliferator-Activated Receptor-γ Agonist, Prevents Microparticle-Induced Vascular Hyporeactivity through the Regulation of Proinflammatory Proteins

Microparticles are plasma membrane vesicles with procoagulant and proinflammatory properties. We recently demonstrated that microparticles induce vascular hyporeactivity and evoke up-regulation of proinflammatory protein expression. This study dissected the effect of either in vitro treatment or short-term oral administration of the peroxisome proliferator-activated receptor-γ (PPARγ) agonist, rosiglitazone, on microparticle-induced vascular hyporeactivity of mouse vessels. Microparticles were produced from T cells by actinomycin D treatment. The effects of rosiglitazone on mouse aortic rings incubated with microparticles were investigated. Aortae treated in vitro with rosiglitazone or aortae taken from mice treated by oral administration of the same agonist completely prevented microparticle-induced vascular hyporeactivity in response to U46619 [9,11-dideoxy-11α, 9α-epoxymethanoprostaglandin F2α). These effects of rosiglitazone occurred independently of the presence of endothelium without modifications in blood parameters. The mechanisms involved abrogation of nitric oxide (NO) and prostacyclin overproduction linked to up-regulation of inducible NO-synthase and cyclooxygenase 2 elicited by microparticles. In addition, rosiglitazone treatment reduced the ability of microparticles to evoke increases in interleukin (IL)-6, IL-8, and nuclear factor (NF)-κB transcription, and NF-κB expression and activation. These results suggest that rosiglitazone, via PPARγ activation, counteracts vascular dysfunction associated with increased release of proinflammatory proteins elicited by microparticles. They underscore therapeutic perspective for rosiglitazone in vascular diseases involving enhanced participation of microparticles

Increase in circulating microparticles in inflammatory bowel disease patients induces vascular alterations

Among the proposed hypothesis for inflammatory bowel disease (IBD) pathogenesis, a vascular cause has been suggested, mostly in reference to anatomic and pathological studies. We previously reported in small mesenteric arteries from Crohn's disease patients a balance between vasoconstrictor products from cyclooxygenase and unidentified vasodilatory products that maintained vascular reactivity in a physiological range despite an increase of circulatory cytokines in these patients (Tabernero et al., Circulation, 2003). Microparticles (MPs) are membrane vesicles released during cell activation and apoptosis. Elevated levels of circulating MPs have been detected in pathologic conditions and their increase has been associated with disease activity and/or prognosis. We investigated whether MPs from IBD patients may play a role in the regulation of endothelial function and vascular reactivity in this disease. Blood samples were obtained either from IBD patients as well as age and sex-matched healthy subjects. MPs concentration and origin were assessed by flow cytometry using specific antibodies. Male Swiss mice were injected intravenously with MPs from IBD patients or with saline solution and sacrified after 24hours. Endothelial function and vascular reactivity were studied on aortic rings and small mesenteric resistance arteries (SMA) by myography and arteriography, respectively. Patients with IBD displayed increased circulating levels of MPs compared to healthy subjects including those from procoagulant, platelet and activated platelet, endothelial, leukocyte and erythrocyte origins. In aorta, MPs from IBD patients compared to saline significantly reduced both endothelium-dependent relaxation to acetylcholine and contraction to serotonin. In SMA, although MPs from IBD patients did not affect flow-induced dilation, a reduced NO-component that was completely compensated by the endothelium-derived hyperpolarizing factor-component of the response was highlighted. Besides, MPs from IBD did not affect myogenic tone in SMA. These results provide further evidence of increased circulating levels of MPs in patients with IBD. MPs origin may play a role in enhanced coagulation and inflammatory states reported in IBD patients. Finally, MPs from IBD patients influence both endothelial dysfunction and vascular hyporeactivity in the experimental model used.(Partially supported by Ferring France Laboratories)

Expression and Biological Activity of Parathyroid Hormone-Related Peptide in Pregnant Rat Uterine Artery: Any Role for 8-Iso-Prostaglandin F2α?

PTHrP is produced in vessels and acts as a local modulator of tone. We recently reported that PTHrP(1–34) is able to induce vasorelaxation in rat uterine arteries, but in pregnancy, this response is blunted and becomes strictly endothelium dependent. The present study aimed to get insights into the mechanisms involved in these changes because the adaptation of uterine blood flow is essential for fetal development. On d 20 of gestation, RT-PCR analysis of uterine arteries showed that PTH/PTHrP receptor (PTH1R) mRNA expression was decreased, whereas that of PTHrP mRNA was increased. This was associated with a redistribution of the PTHrP/PTH1R system, with both PTH1R protein and PTHrP peptide becoming concentrated in the intimal layer of arteries from pregnant rats. On the other hand, the blunted vasorelaxation induced by PTHrP(1–34) in uterine arteries from pregnant rats was specifically restored by indomethacin and a specific cyclooxygenase-2 inhibitor, NS 398. This was associated with an increase in cyclooxygenase-2 expression and in 8-iso-prostaglandin F2α release when uterine arteries from pregnant rats were exposed to high levels of PTHrP(1–34). Most interestingly, 8-iso-prostaglandin F2α itself was able to increase PTHrP expression and reduce PTH1R expression in cultured rat aortic smooth muscle cells. These results suggest a local regulation of uterine artery functions by PTHrP during pregnancy resulting from PTH1R redistribution. Moreover, they shed light on a potential role of 8-iso-prostaglandin F2α