Endothelial Cell-derived Microparticles Research Articles

Translational Research of the Acute Effects of Negative Emotions on Vascular Endothelial Health: Findings From a Randomized Controlled Study.

Provoked anger is associated with an increased risk of cardiovascular disease events. The underlying mechanism linking provoked anger as well as other core negative emotions including anxiety and sadness to cardiovascular disease remain unknown. The study objective was to examine the acute effects of provoked anger, and secondarily, anxiety and sadness on endothelial cell health. Apparently healthy adult participants (n=280) were randomized to an 8-minute anger recall task, a depressed mood recall task, an anxiety recall task, or an emotionally neutral condition. Pre-/post-assessments of endothelial health including endothelium-dependent vasodilation (reactive hyperemia index), circulating endothelial cell-derived microparticles (CD62E+, CD31+/CD42-, and CD31+/Annexin V+) and circulating bone marrow-derived endothelial progenitor cells (CD34+/CD133+/kinase insert domain receptor+ endothelial progenitor cells and CD34+/kinase insert domain receptor+ endothelial progenitor cells) were measured. There was a group×time interaction for the anger versus neutral condition on the change in reactive hyperemia index score from baseline to 40 minutes (P=0.007) with a mean±SD change in reactive hyperemia index score of 0.20±0.67 and 0.50±0.60 in the anger and neutral conditions, respectively. For the change in reactive hyperemia index score, the anxiety versus neutral condition group by time interaction approached but did not reach statistical significance (P=0.054), and the sadness versus neutral condition group by time interaction was not statistically significant (P=0.160). There were no consistent statistically significant group×time interactions for the anger, anxiety, and sadness versus neutral condition on endothelial cell-derived microparticles and endothelial progenitor cells from baseline to 40 minutes. In this randomized controlled experimental study, a brief provocation of anger adversely affected endothelial cell health by impairing endothelium-dependent vasodilation.

Microparticles: potential new contributors to the pathogenesis of systemic sclerosis?

BackgroundMicroparticles (MPs) are membrane-derived vesicles released from cells undergoing activation or apoptosis with diverse proinflammatory and prothrombotic activities, that have been implicated in the pathogenesis of systemic sclerosis (SSc). We aimed to evaluate the plasma levels of platelet-derived microparticles (PMPs), endothelial cell-derived microparticles (EMPs), and monocyte-derived microparticles (MMPs) in SSc patients, and the association between MPs and the clinical features of SSc.MethodsIn this cross-sectional study, 70 patients with SSc and 35 age- and sex-matched healthy controls were evaluated. Clinical and nailfold capillaroscopy (NFC) data were obtained from all patients. Plasma levels of PMPs (CD42+/31+), EMPs (CD105+), and MMPs (CD14+) were quantified by flow cytometry.ResultsPatients were mainly females (90%), with a mean age of 48.9 years old. PMP, EMP, and MMP levels were significantly increased in SSc patients compared to controls (79.2% ± 17.3% vs. 71.0% ± 19.8%, p = 0.033; 43.5% ± 8.7% vs. 37.8% ± 10.4%, p = 0.004; and 3.5% ± 1.3% vs. 1.1% ± 0.5%, p < 0.0001, respectively). PMP levels were significantly higher in patients with positive anti-topoisomerase-I antibodies (p = 0.030) and in patients with a disease duration > 3 years (p = 0.038). EMP levels were lower in patients with a higher modified Rodnan skin score (p = 0.015), and in those with an avascular score > 1.5 in NFC (p = 0.042).ConclusionThe increased levels of PMPs, EMPs and MMPs in scleroderma patients might indicate a possible role for these agents in the pathogenesis of this challenging disease.

Annexin V− and tissue factor+ microparticles as biomarkers for predicting deep vein thrombosis in patients after joint arthroplasty

ObjectiveVenous thromboembolism (VTE) is a common and severe complication of joint arthroplasty. Microparticles (MPs) containing phosphatidylserine (PS) and tissue factor (TF) can trigger coagulation in VTE. This study aims to measure and compare MP levels in joint arthroplasty patients with and without VTE. MethodsThis prospective cohort study enrolled 181 patients who underwent joint arthroplasty. Ultrasound examination was used to diagnose VTE on preoperative day 0 and postoperative day 6. MPs were analysed using transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and flow cytometry. The levels of platelet-derived microparticles (PMPs), endothelial cell-derived microparticles (EMPs), granulocyte-derived microparticles (GMPs), red cell-derived microparticles (RMPs), monocyte-derived microparticles (MMPs), Annexin V+ MPs (AV+ MPs), and tissue factor+ MPs (TF+ MPs) derived from five kinds of MPs were measured on day 0 (before surgery), 1, 2, 3, 4, 5, and 6 after surgery. ResultsThe levels of AV-TF+ EMPs and AV-TF+ MMPs were significantly increased in patients with VTE on postoperative day 5 compared to those without VTE (P = 0.031 and P = 0.031, respectively). ConclusionAV-TF+ MPs may indicate the development of VTE and serve as predictive markers in joint arthroplasty patients.

Characterisation of circulating endothelial microparticles in Behçet's disease: new markers of chronic endothelial damage?

Endothelial cell-derived microparticles (EMPs) are directly indicative of endothelial cell activation or apoptosis, and may also reflect endothelial inflammation, increased coagulation, and vascular tone. The aim of this study is to investigate whether EMPs would be able to evaluate system involvement and be a new indicators of disease activity in Behçet's disease (BD). Thirty-nine consecutive BD patients (who fulfilled the modified 1990 International Study Group on Behçet's disease or the 2006 International Criteria for Behçet's Disease) and 30 age- and sex-matched healthy controls were enrolled. The plasma levels of EMPs were measured by flow cytometry utilising specific labels for endothelial MPs (CD31+ and CD42b-). The levels of circulating EMPs (CD31+ and CD42b-) were significantly elevated in the case group compared with the healthy control group (p =0.000). Moreover, BD patients plasma EMPs were positively correlated with BD current activity form (r=0.802, p =0.000). Vascular and gastrointestinal involvement in BD patients were significantly increased (p=0.004 and p=0.011, respectively) with respect to patients without vascular and gastrointestinal EMPs. Levels of circulating EMPs are elevated in BD patients and correlate with the disease activity; the elevated EMPs may be a potential indicator to predict disease activity of BD. The plasma level of EMPs was increased, which indicated the increased risk of vascular and digestive tract involvement in BD.

Microparticles: An Alternative Explanation to the Behavior of Vascular Antiphospholipid Syndrome.

Antiphospholipid syndrome is an autoimmune disease characterized by the persistent presence of antiphospholipid antibodies, along with occurrence of vascular thrombosis and pregnancy morbidity. The variety of antiphospholipid antibodies and their related mechanisms, as well as the behavior of disease in wide groups of patients, have led some authors to propose a differentiation of this syndrome into two independent entities: vascular and obstetric antiphospholipid syndrome. Thus, previous studies have discussed whether specific autoantibodies may be responsible for this differentiation or, in contrast, how the same antibodies are able to generate two different clinical presentations. This discussion is yet to be settled. The capability of serum IgG from patients with vascular thrombosis to trigger the biogenesis of endothelial cell-derived microparticles in vitro is one of the previously discussed differences between the clinical entities of antiphospholipid syndrome. These vesicles constitute a prothrombotic mechanism as they can directly lead to clot activation in murine models and recalcified human plasma. Nevertheless, other indirect mechanisms by which microparticles can spread a procoagulant phenotype could be critical to understanding their role in antiphospholipid syndrome. For this reason, questions regarding the cargo of microparticles, and the signaling pathways involved in their biogenesis, are of interest in attempting to explain the behavior of this autoimmune disease.

Elevated Circulating Microparticle Subpopulations in Incidental Cerebral White Matter Hyperintensities: A Multimodal Study.

Asymptomatic (or “silent”) manifestations of cerebral small vessel disease (CSVD) are widely recognized through incidental findings of white matter hyperintensities (WMHs) as a result of magnetic resonance imaging (MRI). This study aims to examine the potential associations of surrogate markers for the evaluation of white matter integrity in CSVD among asymptomatic individuals through a battery of profiling involving QRISK2 cardiocerebrovascular risk prediction, neuroimaging, neurocognitive evaluation, and microparticles (MPs) titers. Sixty asymptomatic subjects (mean age: 39.83 ± 11.50 years) with low to moderate QRISK2 scores were recruited and underwent neurocognitive evaluation for memory and cognitive performance, peripheral venous blood collection for enumeration of selected MPs subpopulations, and 3T MRI brain scan with specific diffusion MRI (dMRI) sequences inclusive of diffusion tensor imaging (DTI). WMHs were detected in 20 subjects (33%). Older subjects (mean age: 46.00 ± 12.00 years) had higher WMHs prevalence, associated with higher QRISK2 score and reduced processing speed. They also had significantly higher mean percentage of platelet (CD62P)- and leukocyte (CD62L)-derived MPs. No association was found between reduced white matter integrity—especially at the left superior longitudinal fasciculus (LSLF)—with age and neurocognitive function; however, LSLF was associated with higher QRISK2 score, total MPs, and CD62L- and endothelial cell-derived MPs (CD146). Therefore, this study establishes these multimodal associations as potential surrogate markers for “silent” CSVD manifestations in the well-characterized cardiocerebrovascular demographic of relatively young, neurologically asymptomatic adults. Furthermore, to the best of our knowledge, this study is the first to exhibit elevated MP counts in asymptomatic CSVD (i.e., CD62P and CD62L), which warrants further delineation.

A Novel Pool of Microparticle Cholesterol Is Elevated in Rheumatoid Arthritis but Not in Systemic Lupus Erythematosus Patients

Microparticles are sub-micron, membrane-bound particles released from virtually all cells and which are present in the circulation. In several autoimmune disorders their amount and composition in the circulation is altered. Microparticle surface protein expression has been explored as a differentiating tool in autoimmune disorders where the clinical pictures can overlap. Here, we examine the utility of a novel lipid-based marker—microparticle cholesterol, present in all microparticles regardless of cellular origin—to distinguish between rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). We first isolated a series of microparticle containing lipoprotein deficient fractions from patient and control plasma. There were no significant differences in the size, structure or protein content of microparticles isolated from each group. Compared to controls, both patient groups contained significantly greater amounts of platelet and endothelial cell-derived microparticles. The cholesterol content of microparticle fractions isolated from RA patients was significantly greater than those from either SLE patients or healthy controls. Our data indicate that circulating non-lipoprotein microparticle cholesterol, which may account for 1–2% of measured cholesterol in patient samples, may represent a novel differentiator of disease, which is independent of cellular origin.

Apoptotic endothelial cell-derived microparticles, endothelial progenitor cells and atherosclerosis burden assessed by the friesinger index

Apoptotic endothelial cell-derived microparticles, endothelial progenitor cells and atherosclerosis burden assessed by the friesinger index

Synergistic effects of EMPs and PMPs on pulmonary vascular leakage and lung injury after ischemia/reperfusion

BackgroundVascular leakage is an important pathophysiological process of critical conditions such as shock and ischemia–reperfusion (I/R)-induced lung injury. Microparticles (MPs), including endothelial cell-derived microparticles (EMPs), platelet-derived microparticles (PMPs) and leukocyte-derived microparticles (LMPs), have been shown to participate in many diseases. Whether and which of these MPs take part in pulmonary vascular leakage and lung injury after I/R and whether these MPs have synergistic effect and the underlying mechanism are not known.MethodsUsing hemorrhage/transfusion (Hemo/Trans) and aorta abdominalis occlusion-induced I/R rat models, the role of EMPs, PMPs and LMPs and the mechanisms in pulmonary vascular leakage and lung injury were observed.ResultsThe concentrations of EMPs, PMPs and LMPs were significantly increased after I/R. Intravenous administration of EMPs and PMPs but not LMPs induced pulmonary vascular leakage and lung injury. Furthermore, EMPs induced pulmonary sequestration of platelets and promoted more PMPs production, and played a synergistic effect on pulmonary vascular leakage. MiR-1, miR-155 and miR-542 in EMPs, and miR-126 and miR-29 in PMPs, were significantly increased after hypoxia/reoxygenation (H/R). Of which, inhibition of miR-155 in EMPs and miR-126 in PMPs alleviated the detrimental effects of EMPs and PMPs on vascular barrier function and lung injury. Overexpression of miR-155 in EMPs down-regulated the expression of tight junction related proteins such as ZO-1 and claudin-5, while overexpression of miR-126 up-regulated the expression of caveolin-1 (Cav-1), the trans-cellular transportation related protein such as caveolin-1 (Cav-1). Inhibiting EMPs and PMPs production with blebbistatin (BLE) and amitriptyline (AMI) alleviated I/R induced pulmonary vascular leakage and lung injury.ConclusionsEMPs and PMPs contribute to the pulmonary vascular leakage and lung injury after I/R. EMPs mediate pulmonary sequestration of platelets, producing more PMPs to play synergistic effect. Mechanically, EMPs carrying miR-155 that down-regulates ZO-1 and claudin-5 and PMPs carrying miR-126 that up-regulates Cav-1, synergistically mediate pulmonary vascular leakage and lung injury after I/R.Graphic abstract8k8MRpqBub_wL9wmZ4ez89Video

Association of endothelial and red blood cell microparticles with acute myocardial infarction in Chinese: a retrospective study.

Mounting evidence suggests that endothelial cell-derived microparticles (EMPs) and red blood cell-derived microparticles (RMPs), which have procoagulant and vasoconstriction effects, are involved in the development of vascular acute cardiovascular events. The aim of this study was to analyze the circulating levels of EMPs and RMPs in patients with acute myocardial infarction (AMI), and to explore the correlations between EMPs and RMPs and the severity of coronary artery disease. Plasma samples from 110 patients with AMI and 57 non-coronary artery disease group (nonCAD) were collected in the present study. The flow cytometry was used to measure the EMPs (CD31) and RMPs (CD235a) qualitatively and quantitatively. The levels of EMPs and RMPs in the AMI group were higher than that in the Non-CAD group, yet no significant difference was found between STEMI and non-STEMI subjects. The levels of EMPs and RMPs in multi-vessel were higher than in the single-vessel l disease. In the Thrombolysis in Myocardial Infarction (TIMI) risk assessment, the levels of EMPs in the high-risk group were higher than that in both intermediate- and low-risk group. The low-risk group had the lowest EMP levels, the difference between the groups being statistically significant (P=0.001). No significant difference in RMP levels was noted upon TIMI stratification. According to the ROC curve analysis, the areas under the curve (AUC) of EMPs and RMPs were 0.706 and 0.668, respectively. The circulating levels of EMPs and RMPs in patients with AMI are elevated, and the level of EMPs is related to the degree of coronary artery disease and the prognosis risk. The EMPs are more likely to be potential biomarkers than RMPs to provide diagnostic value for AMI.

Longer Sleep Duration and Endothelial Cell Health Among a Multiethnic Sample of Adolescents.

Adverse endothelial cell health, an early pathogenic process underlying atherosclerosis and cardiovascular disease, is evident in childhood and adolescence. Sleep duration, a modifiable cardiovascular health behavior, may be an important cardiovascular disease prevention target that may affect endothelial cell health. We examined the associations of longer sleep duration with endothelial cell injury among youth. In a multiethnic sample of 235 children (63.0% female, mean age = 13.9 years), we conducted multivariable linear regressions to test the cross-sectional association of sleep duration and circulating levels of endothelial cell-derived microparticles (EMPs), phenotypic for endothelial cell activation and apoptosis (CD62E+ EMPs, CD31+/CD42b- EMPs, and CD31+/Annexin V+ EMPs). Sleep duration and EMPs were both treated as continuous variables. Models were adjusted for age, sex, race, pubertal status, household economic resources, and waist circumference. Overall, 69.2% had short sleep duration (<8 hours of sleep per night). Longer sleep duration was significantly associated with lower levels of CD62E+ EMPs and CD31+/CD42b- EMPs. A 60-minute increase in sleep duration was associated with an 8.40 (95% confidence interval = -205.20 to -1.80, p = .046) decrease in CD62E+ EMPs and a 9.00 (95% confidence interval = -153.60 to -9.60, p = .027) decrease in CD31+/CD42b- EMPs. Sleep duration was not associated with CD31+/Annexin V+ EMPs. Our results support the hypothesis that sleeping longer has beneficial effects on endothelial cell health during childhood. Primordial prevention efforts might incorporate sleep extension to offset cardiovascular risk in youth.

Circulating Endothelial Microparticles Reduce in Concentration Following an Exercise Programme in Women With Polycystic Ovary Syndrome.

Purpose: Endothelial dysfunction is a known comorbidity in women with polycystic ovary syndrome (PCOS). The aim was to assess if supervised, moderate intensity exercise could potentially impact markers of endothelial disruption; endothelial cell derived microparticles (EMP).Methods: The current study investigated the effects of a supervised 8-week moderate intensity exercise programme on EMP in women with PCOS (n = 11) and control women free from any known disease (n = 10). EMP were enumerated via specific antibody (CD105, CD106) labeling and flow cytometry.Results: CD105+MP significantly reduced in women with PCOS from pre to post-exercise programme, with CD105+ MP reducing from 2114 CD105+ MP per μl platelet free plasma (PFP) to 424 CD105+ MP per μl PFP (p = 0.025). Control women showed no significant change in CD105+ MP (p = 0.25) after completing the same exercise programme. CD106+ MP showed no change in either PCOS (p = 0.95) or control groups (p = 0.99). No significant correlations existed with the changes in EMP compared to body composition changes as a result of exercise.Conclusion: Supervised, moderate intensity exercise independent of substantial weight loss reduced circulating CD105+ MP, likely reflecting an improvement in endothelial function in women with PCOS compared to healthy control women. Additionally, EMP may be a useful marker for physical improvement in exercise programmes for clinical populations.

Microparticle Responses to Aerobic Exercise and Meal Consumption in Healthy Men.

Microparticles (MP) are shed extracellular vesicles that express the prothrombotic tissue factor (TF). Aerobic exercise may reduce MP count and TF expression. This study investigated the impact of acute running or rest followed by standardized meal consumption on MP phenotypes and TF expression. Fifteen males (age, 22.9 ± 3.3 yr; body mass, 81.9 ± 11.4 kg; V˙O2max, 54.9 ± 6.5 mL·kg·min; mean ± SD) completed 1 h of running (70% V˙O2max) or rest at 9:00 AM and consumed a standardized meal (1170 kcal, 43% CHO, 17% PRO, 40% fat) at 10:45 AM. Venous blood samples were taken at 9:00 AM, 10:00 AM, and 11:30 AM. The MP concentration, diameter, phenotypes, and TF expression were assessed using nanoparticle tracking analysis and flow cytometry. Nanoparticle tracking analysis identified no changes in MP concentration or diameter in response to time or trial. Flow cytometry revealed total MP count increased from 9:00 AM to 10:00 AM (1.62 ± 2.28 to 1.74 ± 2.61 × 10 L, P = 0.016, effect size (η) = 0.105), but was unaffected by trial. TF platelet-derived MP % reduced from 9:00 AM to 10:00 AM (44.0% ± 21.2% to 21.5% ± 9.3%, P = 0.001, η = 0.582) after exercise only (control, 36.8% ± 18.2% to 34.9% ± 11.9%; P = 0.972). TF neutrophil-derived MP percentage reduced from 9:00 AM to 11:30 AM (42.3% ± 17.2% to 25.1% ± 14.9%; P = 0.048, η = 0.801) in the exercise trial only (control, 28.5% ± 15.7% to 32.2% ± 9.6%; P = 0.508). Running induced a significant reduction in %TF platelet and neutrophil MP, suggesting a transient reduction in cardiovascular risk via reduced TF-stimulated thrombosis. This requires further investigation over longer periods in cardiovascular disease populations.

Microparticles from vascular endothelial growth factor pathway inhibitor-treated cancer patients mediate endothelial cell injury.

Vascular endothelial growth factor pathway inhibitors (VEGFi), used as anti-angiogenic drugs to treat cancer are associated with cardiovascular toxicities through unknown molecular mechanisms. Endothelial cell-derived microparticles (ECMPs) are biomarkers of endothelial injury and are also functionally active since they influence downstream target cell signalling and function. We questioned whether microparticle (MP) status is altered in cancer patients treated with VEGFi and whether they influence endothelial cell function associated with vascular dysfunction. Plasma MPs were isolated from cancer patients before and after treatment with VEGFi (pazopanib, sunitinib, or sorafenib). Human aortic endothelial cells (HAECs) were stimulated with isolated MPs (106 MPs/mL). Microparticle characterization was assessed by flow cytometry. Patients treated with VEGFi had significantly increased levels of plasma ECMP. Endothelial cells exposed to post-VEGFi treatment ECMPs induced an increase in pre-pro-ET-1 mRNA expression, corroborating the increase in endothelin-1 (ET-1) production in HAEC stimulated with vatalanib (VEGFi). Post-VEGFi treatment MPs increased generation of reactive oxygen species in HAEC, effects attenuated by ETA (BQ123) and ETB (BQ788) receptor blockers. VEGFi post-treatment MPs also increased phosphorylation of the inhibitory site of endothelial nitric oxide synthase (eNOS), decreased nitric oxide (NO), and increased ONOO− levels in HAEC, responses inhibited by ETB receptor blockade. Additionally, gene expression of proinflammatory mediators was increased in HAEC exposed to post-treatment MPs, effects inhibited by BQ123 and BQ788. Our findings define novel molecular mechanism involving interplay between microparticles, the ET-1 system and endothelial cell pro-inflammatory and redox signalling, which may be important in cardiovascular toxicity and hypertension associated with VEGFi anti-cancer treatment. New and noteworthy: our novel data identify MPs as biomarkers of VEGFi-induced endothelial injury and important mediators of ET-1-sensitive redox-regulated pro-inflammatory signalling in effector endothelial cells, processes that may contribute to cardiovascular toxicity in VEGFi-treated cancer patients.

Abstract 049: Endothelial-Derived Microparticles as Biomarkers and Mediators of Endothelial Cell Injury in VEGF Inhibitor-Treated Cancer Patients: Implications in Hypertension

Vascular endothelial growth factor receptor inhibitors (VEGFRi), used as anti-angiogenic drugs to treat cancer, induce severe hypertension although the underlying molecular mechanisms are still unclear. Endothelial microparticles (MPs) are biomarkers of endothelial injury and are also functionally active since they influence downstream target cell signalling and function. We questioned whether MP status is altered in cancer patients treated with VEGFRi and whether they influence endothelial cell function associated with vascular dysfunction in hypertension. Plasma MPs were isolated from cancer patients before and after treatment with VEGFRi. Human aortic endothelial cells (HAEC) were stimulated with human plasma-isolated MPs (10 6 MPs/mL). MP characterization was assessed by flow cytometry; protein and gene expression by immunoblotting and qPCR; ROS and NO production by lucigenin and immunofluorescence. Patients treated with VEGFRi had significantly increased in endothelial cell-derived MPs (EMP) (0.19±0.02 Pre vs. 0.31±0.04 Post-treatment). HAEC exposed to post-treatment MPs increased pre-pro-ET-1 mRNA (4.10±0.97 vs. 10.57±3.54), corroborating the raise in ET-1 levels (μg/mL: 631.9±46.1 vs. 780.2±37.2) observed in HAEC stimulated with vatalanib (VEGFRi). Post-treatment MPs increased ROS generation in HAEC (100.0 vs. 5 min- 123.4±7.7, 30 min- 162.4±22.5, 60 min- 183.9±44.7), effects that were attenuated by ET A and ET B receptor blockers. VEGFRi post-treatment MPs increased phosphorylation of the inhibitory site of eNOS (Thr 495 ) (100.0 vs. 181.3±18.7) and decreased NO levels in HAEC (100.0 vs. 72.7±9.9) which was inhibited by ET B receptor blockade (eNOS: 109.8±14.9; NO: 142.1±25.9). Gene expression of proinflammatory mediators was increased in HAEC exposed to post-treatment MPs (1.0 vs. TNF-α: 5.4±1.8, MCP-1: 2.1±0.3, iNOS: 3.1±0.8, COX2: 2.4±0.4, ICAM1: 2.7±0.6), effects inhibited by BQ123 and BQ788. In conclusion, our data identify EMPs as biomarkers of VEGFi-induced endothelial injury and important mediators of ET-1-sensitive redox-regulated endothelial cell signalling. These molecular processes may play a role in vascular dysfunction associated with hypertension in VEGFRi-treated cancer patients.

Putative mechanisms Underlying Myocardial infarction onset and Emotions (PUME): a randomised controlled study protocol

IntroductionThe experience of negative emotions (eg, anger, anxiety and sadness) is associated with an increased short-term risk of incident cardiovascular disease (CVD) events, independent of traditional CVD risk factors. Impairment...

Endothelial microparticles-mediated transfer of microRNA-19b promotes atherosclerosis via activating perivascular adipose tissue inflammation in apoE−/− mice

Microparticles(MPs) are the major carriers of circulating microRNAs. Our previous study has shown that microRNA (miR)-19b in endothelial cell-derived microparticles (EMPs) is significantly increased in patients with unstable angina. However, little is known about the relationship between miR-19b in EMPs and the progression of atherosclerosis. The aim of the present study was to define the role and potential mechanism of miR-19b incorporated in EMPs in the development of atherosclerosis.Western-diet-fed apoE−/− mice were injected with phosphate buffered solution(PBS), EMP carrying microRNA control(EMPcontrol) or miR-19b mimic (EMPmiR19b) intravenously. Systemic treatment with EMPmiR19b significantly accelerated carotid artery atherosclerosis progression by increasing lipid, macrophages and smooth muscle cells and decreasing collagen content in atherosclerotic plaque. Fluorescence-labelled EMPmiR19b injection proved that miR-19b could be transported into perivascular adipose tissue(PVAT) by EMPs. EMPmiR19b treatment also promoted inflammatory cytokines secretion and macrophages infiltration in PVAT. In further experiment, apoE−/− mice were divided into 3 groups: EMPcontrolPVAT(+), EMPmiR19bPVAT(+) and EMPmiR19bPVAT(-), based on removing or keeping pericarotid adipose tissue and injected with EMPcontrol or EMPmiR19b. Loss of PVAT attenuated EMPmiR19b-mediated effects on increasing carotid atherosclerosis formation and inflammatory cytokines level in plaque. EMPmiR19b inhibited suppressor of cytokine signaling 3 (SOCS3) expression in PVAT. Our findings demonstrate that miR-19b in EMPs exaggerates atherosclerosis progression by augmenting PVAT-specific inflammation proceeded by downregulating SOCS3 expression.

Abstract 203: Glucose Stimulated Endothelial Microparticles Increase Endothelial Cell Apoptotic Susceptibility

Clinical interest in endothelial cell-derived microparticles (EMPs) has increased due to their role in the pathogenesis of vascular disease. Although released by the endothelium, EMPs have autocrine properties that can significantly impact endovascular health. Hyperglycemic conditions, such as diabetes, are known to stimulate EMP release; however, the effects of these glucose-related microparticles on endothelial cell function are not well understood. High glucose concentrations induce endothelial cell apoptosis through a caspase-3-dependent mechanism. The aim of this study was to determine the effect of EMPs derived from a hyperglycemic condition on endothelial cell susceptibility to apoptosis. Human umbilical vein endothelial cells (HUVECs) were cultured (3 rd passage) and plated in 6-well plates at a density of 5.0 x 10 5 cell/condition. Cells were incubated with RPMI 1640 media containing 25mM D-glucose (concentration representing a diabetic glycemic state) or 5mM D-glucose (control, normoglycemic, condition) for 48 h to generate EMPs. EMPs derived from both conditions were pelleted by centrifugation and resuspended in culture media. EMP identification (CD144 + expression) and number was determined by flow cytometry. HUVECs (2 x10 6 cells/condition) were treated with EMPs (2:1 ratio) generated from either the hyperglycemic or normoglycemic conditions for 24 h. Thereafter, cells were treated with staurosporine (1μmol/L) for 3 h at 37°C and biotin-ZVKD-fmk inhibitor for 1 h at 37°C. Intracellular concentration of active caspase-3 was determined by enzyme immune assay. Cellular expression of miR-Let7a, an anti-apoptotic microRNA, was determined by RT-PCR using the ΔΔCT normalized to RNU6. Hyperglycemic EMPs resulted in significant increase in basal (1.5 + 0.1 vs 1.0 + 0.1 ng/mL) and staurosporine-stimulated (2.2 + 0.2 vs 1.4 + 0.1 ng/mL) caspase-3 activity compared with normoglycemic EMPs. Additional, the expression of miR-Let7a was markedly reduced (~140%) in response to hyperglycemic EMPs (0.43 + 0.17 fold vs control). These results demonstrate that hyperglycemic-induced EMPs increase endothelial cell apoptotic susceptibility. This apoptotic effect may be mediated, at least in part, by a reduction in miR-Let7a expression.

Sustained apnea induces endothelial activation.

Apnea diving has gained worldwide popularity, even though the pathophysiological consequences of this challenging sport on the human body are poorly investigated and understood. This study aims to assess the influence of sustained apnea in healthy volunteers on circulating microparticles (MPs) and microRNAs (miRs), which are established biomarkers reflecting vascular function. Short intermittent hypoxia due to voluntary breath-holding affects circulating levels of endothelial cell-derived MPs (EMPs) and endothelial cell-derived miRs. Under dry laboratory conditions, 10 trained apneic divers performed maximal breath-hold. Venous blood samples were taken, once before and at 4 defined points in time after apnea. Samples were analyzed for circulating EMPs and endothelial miRs. Average apnea time was 329 seconds (±103), and SpO2 at the end of apnea was 79% (±12). Apnea was associated with a time-dependent increase of circulating endothelial cell-derived EMPs and endothelial miRs. Levels of circulating EMPs in the bloodstream reached a peak 4 hours after the apnea period and returned to baseline levels after 24 hours. Circulating miR-126 levels were elevated at all time points after a single voluntary maximal apnea, whereas miR-26 levels were elevated significantly only after 30 minutes and 4 hours. Also miR-21 and miR-92 levels increased, but did not reach the level of significance. Even a single maximal breath-hold induces acute endothelial activation and should be performed with great caution by subjects with preexisting vascular diseases. Voluntary apnea might be used as a model to simulate changes in endothelial function caused by hypoxia in humans.

Post-Effect of Air Quality Improvement on Biomarkers for Systemic Inflammation and Microparticles in Asthma Patients After the 2008 Beijing Olympic Games: a Pilot Study.

This study's aim was to investigate the post-effect of an air quality improvement on systemic inflammation and circulating microparticles in asthmatic patients during, and 2months after, the Beijing Olympics 2008. We measured the levels of circulating inflammatory cytokines and microparticles in the peripheral blood from asthma patients and healthy controls during (phase 1), and 2months after (phase 2) the Beijing 2008 Olympic Games. The concentrations of circulating cytokines (including TNFα, IL-6, IL-8, and IL-10) were still seen reduced in phase 2 when compared with those in phase 1. The number of circulating endothelial cell-derived microparticles was significantly lower during the phase 2 than that during phase 1 in asthma patients. The level of plasma lipopolysaccharide-binding protein (LBP) was significantly decreased in asthmatics in phase 2. The level of norepinephrine was significantly higher in phase 2 than that in phase 1 in plasma from both asthma patients and healthy subjects. There were no significant differences in the gene profile for the toll-like receptor (TLR) signaling from peripheral blood mononuclear cells. In vitro, microvesicles from patients with asthma impaired the relaxation to bradykinin and contraction to acetylcholine, whereas microparticles from healthy subjects did not. These data suggested that reduction in systemic pro-inflammatory responses and circulating LBP and increased level of norepinephrine in asthma patients persisted even after 2months of the air pollution intervention. These changes were independent of the TLR signaling pathway. Circulating microparticles might be associated with airway smooth muscle dysfunction.