Plasma Endothelial Microparticles Research Articles

Changes and formation mechanism of plasma endothelial microparticles in patients with acute pancreatitis

Changes and formation mechanism of plasma endothelial microparticles in patients with acute pancreatitis

PM2.5 induces pulmonary microvascular injury in COPD via METTL16-mediated m6A modification

Fine particulate matter (PM2.5) exposure is a significant cause of chronic obstructive pulmonary disease (COPD), but the detailed mechanisms involved in COPD remain unclear. In this study, we established PM2.5-induced COPD rat models and showed that PM2.5 induced pulmonary microvascular injury via accelerating vascular endothelial apoptosis, increasing vascular permeability, and reducing angiogenesis, thereby contributing to COPD development. Moreover, microvascular injury in COPD was validated by measurements of plasma endothelial microparticles (EMPs) and serum VEGF in COPD patients. We then performed m6A sequencing, which confirmed that altered N6-methyladenosine (m6A) modification was induced by PM2.5 exposure. The results of a series of experiments demonstrated that the expression of methyltransferase-like protein 16 (METTL16), an m6A regulator, was upregulated in PM2.5-induced COPD rats, while the expression of other regulators did not differ upon PM2.5-induction. To clarify the regulatory effect of METTL16-mediated m6A modification induced by PM2.5 on pulmonary microvascular injury, cell apoptosis, permeability, and tube formation, the m6A level in METTL16-knockdown pulmonary microvascular endothelial cells (PMVECs) was evaluated, and the target genes of METTL16 were identified from a set of the differentially expressed and m6A-methylated genes associated with vascular injury and containing predicted sites of METTL16 methylation. The results showed that Sulfatase 2 (Sulf2) and Cytohesin-1 (Cyth1) containing the predicted METTL16 methylation sites, exhibited higher m6A methylation and were downregulated after PM2.5 exposure. Further studies demonstrated that METTL16 may regulate Sulf2 expression via m6A modification and thereby contribute to PM2.5-induced microvascular injury. These findings not only provide a better understanding of the role played by m6A modification in PM2.5-induced microvascular injury, but also identify a new therapeutic target for COPD.

Association between plasma endothelial microparticles and contrast-induced nephropathy in patients underwent coronary angiography.

We aim to investigate the association between plasma endothelial microparticles (EMPs) and contrast-induced nephropathy of patients underwent coronary angiography.The patients were divided into normal renal function group and renal dysfunction group based on the estimated glomerular filtration rate (eGFR). Among the 180 cases, 117 received determination of EMP and serum creatinine after percutaneous coronary intervention (PCI) and/or coronary angiography. The patients were divided into contrast-induced-nephropathy (CIN) group and non-CIN group. EMPs collection and determination were performed, together with biochemical analysis and digital subtraction angiography (DSA) analysis.Spearman correlation showed that the expression of EMP was negatively correlated with eGFR (r = –0.201, P < .01). The serum hypersensitive C-reactive protein (hs-CRP), cystatin C (Cys-C), uric acid (UA) were significantly higher in CIN group than that in the non CIN group. Spearman correlation showed that the expression of EMP was positively correlated with serum interleukin-6 (IL-6, r = 0.393, P < .01). The expression of EMP was positively correlated with serum hs-CRP (r = 0.360, P < .01). Logistic regression analysis showed that the levels of N-terminal pro-brain natriuretic peptide (NT-proBNP), eGFR, UA, and Cys-C were correlated with the incidence of contrast induced nephropathy.In patients with contrast-induced-nephropathy, the plasma EMPs were significantly increased after coronary angiography. The expression of plasma EMPs may play a role in the occurrence of contrast-induced-nephropathy.

Effects of acute moderate-intensity exercise at different duration on blood pressure and endothelial function in young male patients with stage 1 hypertension

ABSTRACT Purpose The studies regarding the effects of exercise duration on blood pressure have reported inconsistent and conflicting results. Nitric oxide (NO) is a well-known vasodilator released by endothelial cells, and endothelial microparticles (EMPs) are membranous vesicles released into the circulation from activated or apoptotic endothelial cells, both of whose level can reflect the endothelial function. But few studies have been done to explore the effect of exercise duration on blood pressure and endothelial function, especially, the EMPs response to exercise in young male patients with hypertension. This study aimed to investigate the blood pressure response during moderate-intensity exercise performed in different durations and the acute effects on post-exercise hypotension and endothelial function in hypertensive patients. Methods Eighteen young male hypertensive patients who did not take antihypertensive drugs were recruited in this study. They randomly performed twice exercises on a cycle ergometer at a moderate intensity of 40%–50% of their HR reserve; one was 20 min (E20 session), the other one was 40 min (E40 session); there was 1-week break between the two exercises. Blood pressure was monitored by the YUWELL blood pressure monitor at rest, every 5 min during exercise, and 3 and 6 min post-exercise. The level of NO (nitrate/nitrite reduction) and EMPs (flow cytometry) in plasma were detected before and immediately following exercise. CD31+/CD42b− events were classified as EMPs and events per microliter plasma were calculated. This study was approved by the Beijing Sport University Institutional Review Board (protocol number 2019087 H). Results Mean age of patients was 34.8 ± 3.5 yrs, and BMI was 27.6 ± 2.7 kg/m2. Systolic blood pressure (SBP) in the two sessions increased significantly during exercise. The SBP in the E20 session increased by 37 mmHg (P < .01) at 5 min during exercise and remained stable afterward. In the E40 session, the SBP increased by 35 mmHg (P < .01) at 5 min during exercise and was stable at 5 through 25 min, decreased by 8 mmHg at 25 through 35 min during exercise, and then stabilized (P < .01). There was no obvious change of diastolic blood pressure (DBP) in the two sessions during exercise. In the E20 session, SBP at 6 min following exercise was no different from the rest; in the E40 session, SBP at 6 min following exercise was 14 mmHg lower than at the rest (P < .01). DBP in the E20 session was the same as rest at 3 min following exercise and decreased by 4 mmHg at 6 min following exercise compared with the rest (P < .05), while DBP in the E40 session decreased by 3 mmHg at 3 min following exercise compared with the rest (P < .05). The plasma NO levels of E20 and E40 increased significantly following exercise (P < .01 for both). There was no difference in plasma NO level between the two sessions pre- and post-exercise, but the increased magnitude of NO level in E40 was greater than that in E20 (24.47% vs. 9.24%, P < .01). The plasma EMPs level of E20 and E40 decreased significantly following exercise (P < .01 for both). There was no difference in plasma EMPs level between the two sessions pre- and post-exercise, but the decreased magnitude of EMPs level in E40 was greater than that in E20 (15.66% vs. 8.00%, P < .01). Conclusion There is no exaggerated blood pressure response to 20-min and 40-min acute moderate-intensity exercise in young hypertensive men with no antihypertensive drugs. Both 20-min and 40-min acute moderate exercise can reduce the rest blood pressure, and improve endothelial function by increasing NO and decreasing EMPs in young male patients with hypertension, and the effects of 40-min exercise on lowering SBP and improving endothelial function are better than that of 20-min.

Perfusion microvessel density in the cerebral cortex of septic rats is negatively correlated with endothelial microparticles in circulating plasma.

In sepsis, endothelial microparticles (EMPs) released from endothelial cells (ECs) participate in microcirculation dysfunction through pro-coagulant and pro-inflammatory effects, which can lead to sepsis-associated brain dysfunction. However, the relationship between EMPs and cerebral cortical perfusion microvessel density has not been explored. A closed cranial window was created in rats who were tended to until the cerebral cortex edema caused by preparation of the cranial window subsided, and the microvessel density was stable. A cecal ligation and puncture (CLP) sepsis procedure was then performed on day 6, post-surgery. At 12 and 24h after the CLP, cerebral cortical perfusion microvessel density was measured with optical coherence tomography angiography (OCTA), followed by measurement of EMPs to evaluate the relationship between these factors. Microvessel density changed from 46.38 % ± 7.65 % on the day of surgery to 35.87 % ± 11.05 % on the second day and 36.71 % ± 11.38 % on the third day after surgery, and then increased daily. The microvessel density decreased to 27.20 % ± 8.50 % 24h after CLP, which was significantly lower than that immediately and 12h after CLP (P < 0.001). EMPs increased progressively at 12 and 24h after CLP. Moreover, there was a negative correlation between EMPs and microvessel density (r=-0.56, P = 0.01). Edema and microvessel density decreased in the local cerebral cortex of the window and then gradually recovered after cranial window surgery. In sepsis, the perfusion microvessel density of the cerebral cortex negatively correlated with the EMPs. Therefore, the perfusion microvessel density can be indirectly evaluated by detecting the plasma EMP level.

Endothelial Microparticles and Vascular Endothelial Growth Factor in Patients With Head and Neck Cancer Undergoing Radiotherapy or Radiochemotherapy.

Endothelial microparticles (EMPs) released from activated or apoptotic endothelial cells may play a role in coagulation and thrombus formation. However, there is insufficient evidence regarding the impact of EMPs on angiogenesis in patients with cancer. Sixteen patients with head and neck cancer (HNC) undergoing radiotherapy/radiochemotherapy (RT/RCT) and 10 healthy controls were studied. Serum EMPs were counted by flow cytometry, and vascular endothelial growth factor (VEGF) was measured by enzyme-linked immunosorbent assay (ELISA). The mean EMP level was significantly higher in patients with HNC before RT/RCT (1,601±1,479 EMP/μl) compared to the control group (782±698 EMP/μl). The number of EMPs was not notably increased after RT/RCT (1,629±769 EMP/μl). There was no significant correlation between the plasma EMP number and concentration of VEGF before (r=0.131; p=0.625), 1 day after (r=-0.042, p=0.874), nor 3 months after RT/RCT (r=0.454, p=0.076). Released EMPs may not influence promotion of neovascularization in patients with HNC.

Altered lung biology of healthy never smokers following acute inhalation of E-cigarettes

BackgroundLittle is known about health risks associated with electronic cigarette (EC) use although EC are rising in popularity and have been advocated as a means to quit smoking cigarettes.MethodsTen never-smokers, without exposure history to tobacco products or EC, were assessed at baseline with questionnaire, chest X-ray, lung function, plasma levels of endothelial microparticles (EMP), and bronchoscopy to obtain small airway epithelium (SAE) and alveolar macrophages (AM). One week later, subjects inhaled 10 puffs of “Blu” brand EC, waited 30 min, then another 10 puff; n = 7 were randomized to EC with nicotine and n = 3 to EC without nicotine to assess biological responses in healthy, naive individuals.ResultsTwo hr. post-EC exposure, subjects were again assessed as at baseline. No significant changes in clinical parameters were observed. Biological changes were observed compared to baseline, including altered transcriptomes of SAE and AM for all subjects and elevated plasma EMP levels following inhalation of EC with nicotine.ConclusionsThis study provides in vivo human data demonstrating that acute inhalation of EC aerosols dysregulates normal human lung homeostasis in a limited cohort of healthy naïve individuals. These observations have implications to new EC users, nonsmokers exposed to secondhand EC aerosols and cigarette smokers using EC to quit smoking.Trial registrationClinicalTrials.gov NCT01776398 (registered 10/12/12), NCT02188511 (registered 7/2/14).

High-Mobility Group Box 1 From Hypoxic Trophoblasts Promotes Endothelial Microparticle Production and Thrombophilia in Preeclampsia.

Thrombophilia is a major complication in preeclampsia, a disease associated with placental hypoxia and trophoblast inflammation. Preeclampsia women are known to have increased circulating microparticles that are procoagulant, but the underlying mechanisms remain unclear. In this study, we sought to understand the mechanism connecting placental hypoxia, circulating microparticles, and thrombophilia. We analyzed protein markers on plasma microparticles from preeclampsia women and found that the increased circulating microparticles were mostly from endothelial cells. In proteomic studies, we identified HMGB1 (high-mobility group box 1), a proinflammatory protein, as a key factor from hypoxic trophoblasts in stimulating microparticle production in human umbilical vein endothelial cells. Immunodepletion or inhibition of HMGB1 in the conditioned medium from hypoxic human trophoblasts abolished the endothelial microparticle-stimulating activity. Conversely, recombinant HMGB1 stimulated microparticle production in cultured human umbilical vein endothelial cells. The microparticles from recombinant HMGB1-stimulated human umbilical vein endothelial cells promoted blood coagulation and neutrophil activation in vitro. Injection of recombinant HMGB1 in pregnant mice increased plasma endothelial microparticles and promoted blood coagulation. In preeclampsia women, elevated placental HMGB1 expression was detected and high levels of plasma HMGB1 correlated with increased plasma endothelial microparticles. Our results indicate that placental hypoxia-induced HMGB1 expression and release from trophoblasts are important mechanism underlying increased circulating endothelial microparticles and thrombophilia in preeclampsia.

Association between vitamin D and vascular endothelial injury in hypertensive patients

Objective To evaluate the association between vitamin D (VD) and vascular endothelial injury in hypertensive patients. Methods Ninety nine patients with hypertension and 126 healthy subjects (control group) from the same community in Nanjing were recruited in the study. The serum levels of fasting blood glucose (GLU), total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C) and triglycerides (TG) were measured. Serum 25-hydroxy vitamin D (25-OH-D) and nitric oxide (NO) levels were determined by enzyme-linked immunosorbent assay. The level of plasma endothelial microparticles was measured by flow cytometry. The data were statistically analyzed with SPSS version 19.0. Results The serum 25-(OH)-D[(55.22±11.92)nmol/L vs. (64.17±21.52) nmol/L, t=-3.71], vitamin D binding protein (VDBP)[(257.32±141.31) ng/ml vs. (314.21±124.13) ng/ml, t=-3.21], NO levels [(39.35±13.33) μmol/L vs. (43.42±11.83) μmol/L, t=-2.42]in hypertension group were significantly lower than those in control group. And the circulating endothelial cell particles(4.95±1.78 vs. 2.84±2.07, t=8.05)in hypertension group was significantly higher than that in control group. In hypertensive patients, the circulating endothelial cell particles in VD deficiency group was significantly higher than that in low VD group and normal VD group [(6.42±1.25)% vs. (4.6±1.68)%, (3.15±0.90)%, F=15.08]. And the serum NO level in VD deficiency group was significantly lower than that in other two groups[(36.57±10.52) μmol/L vs. (39.77±13.01) μmol/L, (46.26±25.90) mol/L, F=1.20]. Multiple linear regression analysis showed that low serum 25 (OH) D(B=-0.189, SE=0.033, β=0.509, t=-5.72)and gender (B=-0.682, SE=0.351, β=-0.182, t=-2.054)were the risk factors of vascular endothelial injury. Conclusion The serum vitamin D levels in hypertensive patients is decreased, and vitamin D deficiency may be a risk factor for endothelial injury in hypertension. Key words: Vitamin D; Hypertension; Vascular endothelial cells injury

Endothelial microparticles are increased in congenital heart diseases and contribute to endothelial dysfunction

BackgroundWe previously demonstrated that endothelial microparticles (EMPs) are increased in mitral valve diseases and impair valvular endothelial cell function. Perioperative systemic inflammation is an important risk factor and complication of cardiac surgery. In this study, we investigate whether EMPs increase in congenital heart diseases to promote inflammation and endothelial dysfunction.MethodsThe level of plasma EMPs in 20 patients with atrial septal defect (ASD), 23 patients with ventricular septal defect (VSD), and 30 healthy subjects were analyzed by flow cytometry. EMPs generated from human umbilical vascular endothelial cells (HUVECs) were injected into C57BL6 mice, or cultured with HUVECs without or with siRNAs targeting P38 MAPK. The expression and/or phosphorylation of endothelial nitric oxide synthase (eNOS), P38 MAPK, and caveolin-1 in mouse heart and/or in cultured HUVECs were determined. We evaluated generation of nitric oxide (NO) in mouse hearts, and levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in cultured HUVECs and in mice.ResultsEMPs were significantly elevated in patients with ASD and VSD, especially in those with pulmonary hypertension when compared with controls. EMPs increased caveolin-1 expression and P38 MAPK phosphorylation and decreased eNOS phosphorylation and NO production in mouse hearts. EMPs stimulated P38 MAPK expression, TNF-α and IL-6 production, which were all inhibited by siRNAs targeting P38 MAPK in cultured HUVECs.ConclusionsEMPs were increased in adult patients with congenital heart diseases and may contribute to increased inflammation leading to endothelial dysfunction via P38 MAPK-dependent pathways. This novel data provides a potential therapeutic target to address important complications of surgery of congenial heart disease.

Plasma endothelial microparticles reflect the extent of capillaroscopic alterations and correlate with the severity of skin involvement in systemic sclerosis

Introduction and aimEndothelial microparticles (EMPs) are membrane-coated vesicles shed from endothelial cells and are considered markers of the endothelial state. It has been shown that total numbers of circulating EMPs are increased in patients with systemic sclerosis (SSc), but their clinical correlations have not yet been investigated in detail. We aimed to assess possible relationships between circulating EMPs and clinical as well as laboratory features among SSc patients with special attention to possible association with alteration in microvascular morphology objectified on nailfold videocapillaroscopy and clinical signs of microvascular complications. Materials and methodsThe study included 47 SSc patients and 27 age- and sex-matched healthy controls. EMPs were identified with flow cytometry after staining platelet-poor plasma with combinations of fluorescent cell-specific monoclonal antibodies (anti-CD31, -51, -42b, -62E and Annexin V). The following types of EMPs were evaluated: total EMPs (CD31+/CD42b−), activated EMPs (CD62E+/AnnV−,) and apoptotic EMPs (CD62E+/AnnV+ or CD51+). Clinical evaluation of patients was obtained, including nailfold videocapillaroscopy. ResultsAll types of EMPs were significantly elevated in SSc patients as compared with healthy controls. We found significant inverse correlation between severity of skin involvement and values of total EMPs (r=−0.32; p=0.02) and their levels tended to be lower in SSc patients with digital ulcers when compared to those without ischaemic skin lesions (p=0.09). Total EMPs and activated EMPs showed correlations with the number of ramified capillaries (r=−0.40 and r=0.37, respectively, p<0.05 for both). Moreover, total EMPs inversely correlated with the severity of capillary loss (r=−0.35, p<0.05) and their levels were significantly lower in patients with late NVC pattern with respect to those with early microangiopathy (p<0.05). On the other hand, active NVC pattern was characterized by strongly elevated levels of activated EMPs when compared to an early vascular alteration (p<0.05). ConclusionsOur results suggest that quantity and phenotype of circulating EMPs might indicate on molecular vascular damage with endothelial dysfunction and to reflect progressive loss of capillaries consequencing in microvascular insufficiency in SSc patients.

Endothelial microparticles carrying hedgehog-interacting protein induce continuous endothelial damage in the pathogenesis of acute graft-versus-host disease.

Accumulating evidence suggests that endothelial microparticles (EMPs), a marker of endothelial damage, are elevated in acute graft-versus-host disease (aGVHD), and that endothelial damage is implicated in the pathogenesis of aGVHD, but the mechanisms remain elusive. In this study, we detected the plasma EMP levels and endothelial damage in patients and mice with aGVHD in vivo and then examined the effects of EMPs derived from injured endothelial cells (ECs) on endothelial damage and the role of hedgehog-interacting protein (HHIP) carried by EMPs in these effects in vitro. Our results showed that EMPs were persistently increased in the early posttransplantation phase in patients and mice with aGVHD. Meanwhile, endothelial damage was continuous in aGVHD mice, but was temporary in non-aGVHD mice after transplantation. In vitro, EMPs induced endothelial damage, including increased EC apoptosis, enhanced reactive oxygen species, decreased nitric oxide production and impaired angiogenic activity. Enhanced expression of HHIP, an antagonist for the Sonic hedgehog (SHH) signaling pathway, was observed in patients and mice with aGVHD and EMPs from injured ECs. The endothelial damage induced by EMPs was reversed when the HHIP incorporated into EMPs was silenced with an HHIP small interfering RNA or inhibited with the SHH pathway agonist, Smoothened agonist. This work supports a feasible vicious cycle in which EMPs generated during endothelial injury, in turn, aggravate endothelial damage by carrying HHIP into target ECs, contributing to the continuously deteriorating endothelial damage in the development of aGVHD. EMPs harboring HHIP would represent a potential therapeutic target for aGVHD.

Effects of Chemotherapy and Radiation Exposure on Brachial Artery Blood Flow during Dynamic Handgrip Exercise

Chemotherapy and radiation cancer treatments have been shown to alter peripheral cardiovascular function. Therefore, the purpose of this study was to investigate if the vascular responses to dynamic handgrip exercise are blunted in individuals who have undergone chemotherapy and/or radiation treatment. To date, 1 cancer survivor (CS) and 1 healthy control (HC) have completed two dynamic handgrip exercise protocols at relative (10 and 20% maximal voluntary contraction (MVC)) and absolute (3 and 9 kg) workloads. Simultaneous measurements of forearm blood flow (FBF; Doppler Ultrasound), and mean arterial pressure (MAP) were taken during each of the workloads. Forearm vascular conductance (FVC) was calculated from MAP and FBF. Plasma concentrations of endothelial microparticles (EMPs) which are suggestive of apoptotic and activated endothelial cells (CD31+/CD42b‐ and CD62E+ EMPs, respectively) were isolated from venous blood samples. Preliminary results: FBF was decreased in CS compared to HC at: 10% MVC (111.72 vs. 228.17 ml min‐1) and 20% MVC (148.60 vs. 362.53 ml min‐1). FVC was reduced in CS when compared to HC at: 10% MVC (107.65 vs. 205.32 ml min‐1 100 mmHg‐1) and 20% MVC (135.35 vs. 287.64 ml min‐1 100 mmHg‐1). During absolute workloads FBF and FVC of the CS compared to HC was similar at 3 kg (143.91 vs. 135.26 ml min‐1 and 162.33 vs. 124.42 ml min‐1 100 mmHg‐1, respectively), but less at 9 kg (156.11 vs. 307.42 ml min‐1 and 155.24 vs. 263.54 ml min‐1 100 mmHg‐1, respectively). Due to time constraints plasma EMP concentrations are unavailable at this time. In conclusion, chemotherapy and radiation treatment in cancer survivors results in alterations to exercise blood flow regulation during dynamic exercise.

MicroRNA-19b functions as potential anti-thrombotic protector in patients with unstable angina by targeting tissue factor

The activation of a hemostatic system plays a critical role in the incidence of acute coronary events. Hemostatic proteins may be regulated by microRNAs (miRNAs). Microparticles (MPs) are the major carrier of circulating miRNAs. The aim of this study was to determine the potential role of miRNAs in regulating gene expression involved in the hemostatic system in patients with unstable angina (UA). MiRNA expression profiles in the plasma from patients with UA (UA group, n=9) compared with individuals with clinical suspicion of coronary artery disease (CAD) but negative angiography (control group, n=9) showed that among 36 differentially expressed miRNAs, miR-19b was the most obvious one. Using real-time PCR, 5 selected miRNA levels in plasma (UA group, n=20; control group, n=30) and plasma MPs (UA group n=6; control group n=6) were proved to be consistent with the miRNA array. Flow cytometry analysis indicated that the amounts of plasma endothelial microparticles (EMPs) were increased in UA patients (UA group, n=4) compared to controls (control group, n=4). In cultured endothelial cells (ECs), TNF-α increased miR-19b release and expression. Tissue factor (TF) was predicted to be the target of miR-19b by bioinformatics analysis. Luciferase reporter assays demonstrated that miR-19b binds to TF mRNA. Overexpression of miR-19b inhibited TF expression and procoagulant activity. This study indicates that in UA patients, the increase of miR-19b wrapped in EMPs due to endothelial dysfunction may partially contribute to the circulating miR-19b elevation and miR-19b may play an anti-thrombotic role by inhibiting the expression of TF in ECs.

Circulating endothelial microparticles involved in lung function decline in a rat exposed in cigarette smoke maybe from apoptotic pulmonary capillary endothelial cells.

Plasma levels of endothelial microparticles (EMPs), small membrane vesicles, shed from activated or apoptotic endothelial cells are elevated in patients with COPD and in smokers with normal lung function. Whether plasma EMPs levels are elevated in a rat exposed in cigarette smoke, whether the elevated EMPs derived from pulmonary endothelial cell apoptosis, and the relationship between EMP and lung function are obscure. All 60 wister rats were divided into six groups, three groups of ten rats were exposed to cigarette smoke of ten non-filter cigarettes per day, 5 days a week, using a standard smoking machine (Beijing BeiLanBo Company, China) for a period of 2, 4 and 6 months (n=10, respectively). Age-matched three control groups were sham-smoked. Pulmonary function parameters, including the ratio of forced expiratory volume in 0.3 second over forced vital capacity (FEV0.3/FVC) and dynamic compliance (Cdyn), were tested at the end of each period (2, 4, 6 months). Blood samples were collected and platelet-free plasma was isolated. Then CD42b-/CD31+ EMPs were analysed by flow cytometry. In parallel, lungs were removed and Colocalization with terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL), Hoeschts and CD31 was performed to evaluate pulmonary capillaries-specific apoptosis and identify the origins of the EMPs. At 2, 4 and 6 months, in comparison with control groups, rats in cigarette smoke exposed groups had a significant increase in CD42b-/CD31+ EMPs (P<0.001, P<0.001, P<0.001, respectively), and Pulmonary function indicated that FEV0.3/FVC (P<0.05, P<0.01, P<0.01, respectively) and Cdyn (P<0.01, P<0.001, P<0.001 respectively) decreased. At the same time, CD42b-/CD31+ EMP counts were negatively correlated with Cdyn (P<0.05). Moreover, in vivo, TUNEL-positive cells co-localized with CD31 in whole lung tissue demonstrated a sequence of apoptosis signal in the cigarette smoke exposed groups. CD42b-/CD31+ EMPs may be a potential biomarker for indicating the severity of impairment of pulmonary function in the rats exposed cigarette smoke. The increased EMPs may derive from pulmonary capillaries-specific apoptosis.

Effects of moderate aerobic exercise training on vascular health and blood pressure in African Americans.

As healthcare progresses toward individualized medicine, understanding how different racial groups respond to lifestyle interventions is valuable. It is established that African Americans have disproportionate levels of cardiovascular disease and impaired vascular health, and clinical practice guidelines suggest lifestyle interventions as the first line of treatment. Recently, the authors reported that 6months of aerobic exercise improved inflammatory markers, flow-mediated dilation (FMD), and levels of circulating endothelial microparticles (EMPs) in African American adults. This study is a subgroup analysis of the aerobic exercise-induced changes in vascular health and blood pressure (BP) measures, including carotid artery intima-media thickness (IMT), nitroglycerin-mediated dilation (NMD), ambulatory BP, and office BP. Sedentary African American adults (53.4±6.2years; 21 women and 5 men) showed improved vascular health but no change in BP. Carotid artery IMT decreased 6.4%, plasma nitric oxide levels increased 76.6%, plasma EMP levels decreased, percentage of FMD increased 59.6%, and FMD/NMD ratio increased 36.2% (P<.05 for all). Six months of aerobic exercise training is sufficient to elicit improvements in vascular structure and function in African Americans, even without improvements in BP measures or NMD (ie, smooth muscle function). To our knowledge, this is the first study to report such findings in African Americans.

Evaluation of plasma endothelial microparticles in pre-eclampsia

To compare the antepartum and postpartum plasma concentrations of endothelial microparticles (EMPs) in patients with pre-eclampsia with those of healthy control subjects with normal pregnancies. Plasma concentrations of EMPs were measured using specific antibody labelling and flow cytometry prior to delivery by caesarean section (antepartum), and again at 24 h and 72 h postpartum. The correlations between EMP concentration and mean arterial pressure (MAP), and between EMP concentration and 24-h urine protein were analysed. A total of 59 patients with pre-eclampsia and 60 healthy control subjects participated in the study. The antepartum plasma EMP concentration was significantly higher in patients with pre-eclampsia than in healthy control subjects with normal pregnancies (2863.5 ± 1543.1 versus 1703.0 ± 592.2 EMPs/ml, respectively); similar findings were observed at 24 h postpartum (1836.7 ± 1153.8 versus 1421.8 ± 760.2 EMPs/ml, respectively). There was no significant difference between the two groups at 72 h postpartum. The antepartum plasma EMP concentration in patients with pre-eclampsia demonstrated a significant positive correlation with MAP (r = 0.716) and with 24-h urine protein (r = 0.770). Plasma EMP concentration might serve as a biomarker to evaluate the severity of pre-eclampsia in the future.

Endothelial microparticles increase in mitral valve disease and impair mitral valve endothelial function

Mitral valve endothelial cells are important for maintaining lifelong mitral valve integrity and function. Plasma endothelial microparticles (EMPs) increased in various pathological conditions related to activation of endothelial cells. However, whether EMPs will increase in mitral valve disease and their relationship remains unclear. Here, 81 patients with mitral valve disease and 45 healthy subjects were analyzed for the generation of EMPs by flow cytometry. Human mitral valve endothelial cells (HMVECs) were treated with EMPs. The phosphorylation of Akt and endothelial nitric oxide synthase (eNOS), the association of eNOS and heat shock protein 90 (HSP90), and the generation of nitric oxide (NO) and superoxide anion (O(2)(∙-)) were measured. EMPs were increased significantly in patients with mitral valve disease compared with those in healthy subjects. EMPs were negatively correlated with mitral valve area in patients with isolated mitral stenosis. EMPs were significantly higher in the group with severe mitral regurgitation than those in the group with mild and moderate mitral regurgitation. Furthermore, EMPs were decreased dramatically in both Akt and eNOS phosphorylation and the association of HSP90 with eNOS in HMVECs. EMPs decreased NO production but increased O(2)(∙-) generation in HMVECs. Our data demonstrated that EMPs were significantly increased in patients with mitral valve disease. The increase of EMPs can in turn impair HMVEC function by inhibiting the Akt/eNOS-HSP90 signaling pathway. These findings suggest that EMPs may be a therapeutic target for mitral valve disease.

Increased Carotid Intima-Media Thickness and Reduced Distensibility in Human Class III Obesity: Independent and Differential Influences of Adiposity and Blood Pressure on the Vasculature

Carotid intima-media-thickness (cIMT) and carotid distensibility (distensibility), structural and functional properties of carotid arteries respectively, are early markers, as well as strong predictors of cardiovascular disease (CVD). The characteristic of these two parameters in individuals with BMI>40.0 kg/m2 (Class III obesity), however, are largely unknown. The present study was designed to document cIMT and distensibility in this population and to relate these to other factors with established association with CVD in obesity. The study included 96 subjects (65 with BMI>40.0 kg/m2 and 31, age- and gender-matched, with BMI of 18.5 to 30.0 kg/m2). cIMT and distensibility were measured by non-invasive high resolution ultrasonography, circulatory CD133+/KDR+ angiogenic cells and endothelial microparticles (EMP) by flow cytometry, and plasma levels of adipokines, growth factors and cytokines by Luminex immunoassay kits. The study results demonstrated increased cIMT (0.62±0.11 mm vs. 0.54±0.08 mm, P = 0.0002) and reduced distensibility (22.52±10.79 10−3kpa−1 vs. 29.91±12.37 10−3kpa−1, P<0.05) in individuals with BMI>40.0 kg/m2. Both cIMT and distensibility were significantly associated with traditional CVD risk factors, adiposity/adipokines and inflammatory markers but had no association with circulating angiogenic cells. We also demonstrated, for the first time, elevated plasma EMP levels in individuals with BMI>40.0 kg/m2. In conclusion, cIMT is increased and distensibility reduced in Class III obesity with the changes predominantly related to conventional CVD risk factors present in this condition, demonstrating that both cIMT and distensibility remain as CVD markers in Class III obesity.

Abstract 208: Laminar Shear Stress Decreases Endothelial Microparticles Release by a Nitric Oxide- and ABCA1-Dependent Mechanism

Microparticles (MP) are shed-membrane sub-micron vesicles released upon cell activation or apoptosis. Their formation is associated with loss of cell membrane asymmetry and exposure of phosphatidylserine (PS), resulting from increases in floppase activities such as ABCA1. Plasma endothelial MP (EMP) are surrogate markers of endothelial dysfunction and their levels increase in patients with cardiovascular diseases. However, the mechanisms governing EMP release in vivo remain largely unknown. As nitric oxide (NO) is a key regulator of the vasoprotective effect of laminar shear stress (SS) on the endothelium, we tested the hypothesis that endothelial NO regulates EMP release under different SS profiles. Shed-annexinV+ EMPs were analyzed by flow cytometry in the medium of confluent HUVECs (passage 1-3) subjected to either high- (15 dyne/cm2; atheroprotective) or low- (1.5 dyne/cm2; atheroprone) SS conditions for 24 hrs. HUVECs exposed to high-SS released 3-fold less EMPs compared to low-SS conditions (326±37 vs. 130±23AnnV+MPs/μL, p&lt;0.001). High-SS EMPs also externalized less PS, as estimated by the mean fluorescence of FITC-AnnV labelling (15.6±6.1 vs. 24.2±9.4 AU, p&lt;0.001). Exposure to L-NAME (10-5M), but not D-NAME, increased high-SS-induced EMP release by over 3-fold (p&lt;0.001) and the NO donor SNAP (10-5M) significantly decreased low-SS-induced EMP formation by 50% (p=0.05). In addition, L-NAME increased the mean fluorescence of EMP labelling under high SS (p&lt;0.05), whereas SNAP decreased it under low SS (p&lt;0.05). We then evaluated ABCA1 expression under these conditions. high-SS significantly decreased the expression of ABCA1 both at the mRNA and protein level by 2- and 20-fold, respectively (p&lt;0.05 vs low SS). L-NAME augmented ABCA1 mRNA level in cells exposed to high SS (p&lt;0.05), whereas SNAP significantly decreased ABCA1 mRNA level under low SS conditions (p&lt;0.05). Finally, the LXRα agonist TO90131 (0.1mM) not only increased endothelial ABCA1 expression by 20-fold, but also augmented MP level under high SS conditions. Altogether, these results demonstrate that atheroprotective shear stress limits the release of procoagulant endothelial MP through a NO-dependent regulation of endothelial ABCA1 expression.