Apoptotic Endothelial Microparticles Research Articles

Biomarkers of endothelial activation and dysfunction in cardiovascular diseases.

Endothelial activation and dysfunction is an important contributor to atherosclerosis, cardiovascular diseases and cardiorenal syndrome. Endothelial dysfunction is also linked with metabolic syndrome and type II diabetes. The search for specific and sensitive biomarkers of endothelial activation and dysfunction may have important clinical implications. This review pinpoints the differences in biomarkers between endothelial activation and endothelial dysfunction in cardiovascular diseases, and then briefly describes the most relevant biomarkers of endothelial activation. Biomarkers of endothelial activation include endothelial adhesion molecules, cytokines, C-reactive protein, CD62E+/E-selectin activated endothelial microparticles, oxidation of low density lipoproteins, asymmetric dimethylarginine and endocan. This review also presents an update on the novel biomarkers of endothelial dysfunction, such as matrix metalloproteinases (e.g., MMP-7, MMP-9), ANGPTL2, endogdlin, annexin V+ endothelial apoptotic microparticles, and serum homocysteine. Finally, this review emphasizes the limitations of biomarkers of endothelial activation and dysfunction in clinical setting.

Identification of Fibrinogen as a Key Anti-Apoptotic Factor in Human Fresh Frozen Plasma for Protecting Endothelial Cells In Vitro.

Resuscitation with human fresh frozen plasma (FFP) in hemorrhagic shock (HS) patients is associated with improved clinical outcomes. Our group has demonstrated that the beneficial effect of FFP is due to its blockade on endothelial hyperpermeability, thereby improving vascular barrier function. The current study aimed to investigate HS-induced endothelial cell apoptosis, a potential major contributor to the endothelial hyperpermeability, and to determine the effect and the key components/factors of FFP on protecting endothelial cells from apoptosis. We first measured and demonstrated an increase in apoptotic endothelial microparticles (CD146AnnexinV) in patients in shock compared to normal subjects, indicating the induction of endothelial cell activation and apoptosis in shock patients. We then transfused HS rats with FFP and showed that FFP blocked HS-induced endothelial cell apoptosis in gut tissue. To identify the anti-apoptotic factors in FFP, we utilized high-performance liquid chromatography, fractionated FFP, and screened the fractions in vitro for the anti-apoptotic effects. We selected the most effective fractions, performed mass spectrometry, and identified fibrinogen as a potent anti-apoptotic factor. Taken together, our findings suggest that HS-induced endothelial apoptosis may constitute a major mechanism underlying the vascular hyperpermeability. Furthermore, the identified anti-apoptotic factor fibrinogen may contribute to the beneficial effects of FFP resuscitation, and therefore, may have therapeutic potential for HS.

Pseudoexfoliative Glaucoma, Endothelial Dysfunction, and Arterial Stiffness: The Role of Circulating Apoptotic Endothelial Microparticles.

PExG is characterized by the deposition of pseudoexfoliative material in several tissues and organs including in the cardiovascular system. This study aimed to evaluate the vascular endothelial function, arterial stiffness, inflammatory status, and circulating microparticle (MP) levels in PExG patients compared with those in primary open-angle glaucoma (POAG) patients and control subjects. Vascular endothelial function was evaluated by flow-mediated dilation. Pulse wave velocity and augmentation index were measured as indices of aortic stiffness and arterial wave reflections, respectively. Growth-differentiation factor-15 and intercellular adhesion molecule1 levels were measured to evaluate the systemic inflammatory status. Circulating MPs that constitute an emerging marker of vascular endothelial dysfunction and platelet activation were isolated and analyzed by flow cytometry. There was a stepwise impairment from the control to the POAG patients and PExG subjects in the flow-mediated dilation (8.21%±2.94% vs. 7.56%±3.12% vs. 5.79±3.13, P=0.005), pulse wave velocity (8.14±1.79 vs. 9.21±2.27 vs. 9.95±3.28 m/s, P=0.007), augmentation index (24.71%±7.84% vs. 26.78%±7.21% vs. 29.96%±7.58%, P=0.02), and growth-differentiation factor-15 (P=0.001) and intercellular adhesion molecule1 levels (P=0.08). PExG patients expressed greater levels of total circulating MPs (Annexin V+) (P=0.004) and endothelial-derived MPs (CD144+) (P<0.001) compared with POAG and control subjects. PExG patients with an accumulation of pseudoexfoliative microfibrillar material presented with vascular endothelial dysfunction and arterial wall impairment associated with the levels of circulating proinflammatory molecules and circulating apoptotic endothelial MPs. These findings highlight the underlying systemic pathophysiological mechanisms associated with the progress of the pseudoexfoliative syndrome.

Anti-endothelial cell antibodies are associated with apoptotic endothelial microparticles, endothelial sloughing and decrease in angiogenic progenitors in systemic sclerosis.

IntroductionEvidence has accumulated for the role of endothelial damage in systemic sclerosis (SSc) and the anti-endothelial cell antibodies (AECAs) might underlie vascular injury.AimSince endothelial microparticles (EMPs) and circulating endothelial cells (CECs) reflect endothelial damage, we aimed to investigate their possible relationship with AECAs in SSc. We examined whether AECAs could affect endothelial repair based on the number of endothelial progenitor cells (EPCs).Material and methodsForty-seven SSc patients were screened. The AECAs were identified in serum by indirect immunofluorescence. EPCs and CECs were isolated from the peripheral blood using anti-CD34-based immunomagnetic separation, whereas EMPs were analyzed in plasma. Flow cytometry was used to quantify EMPs, CECs and EPCs.ResultsAECAs were found in 21 (44.7%) SSc patients and were significantly associated with higher levels of total as well as apoptotic (AnnV+ and CD51+) EMPs, whereas activated (CD62E+/AnnV–) EMPs did not differ between groups. Patients with AECAs had significantly elevated total CECs as well as activated CD105+ CECs. Total endothelial progenitors did not differ between patients with or without AECAs; however AECAs was negatively associated with the population of EPCs that express VEGFR2 or Tie2 receptors.ConclusionsWe found an association between AECAs and the severity of endothelial damage in SSc based on higher levels of total EMPs and CECs. In our study, AECAs were associated with apoptosis of ECs rather than their activation. We also identified a possible role of AECAs in the impairment of vascular repair in SSc as evidenced by significantly fewer angiogenic EPCs.

Moderate-intensity exercise reduces activated and apoptotic endothelial microparticles in healthy midlife women.

Endothelial microparticles (EMPs) are related to cardiovascular disease (CVD) risk. Risk factors for CVD increase with menopause, and greater cardiorespiratory fitness is generally expected to reduce CVD risk. The effects of habitual physical activity on endothelial health may be due in part to the effect of acute exercise on circulating EMPs. This study was performed to evaluate the effect of an acute bout of exercise on CD62E+ and CD31+/42b- EMPs in healthy fit midlife women at different menopausal stages. Healthy, active premenopausal (PRE), perimenopausal (PERI), and postmenopausal (POST) women completed a single bout of moderate-intensity treadmill exercise. Activated (CD62E+) and apoptotic (CD31+/42b-) EMPs were evaluated before and 30 min after exercise by using fluorescent activated cell sorting. In an exploratory analysis, these results were compared with data from low-fit peri- and postmenopausal women. Differences by group and time point were evaluated with repeated-measure ANOVAs. There was a reduction in the number of total microparticles ( P < 0.001), CD62E+ ( P = 0.003), and CD31+/42b- ( P < 0.001) EMPs/μl plasma following acute exercise. The percentage of CD62E+ EMPs increased with acute exercise ( P < 0.001), whereas the percentage of CD31+/42b- EMPs did not change ( P = 0.40). There was no effect of menopausal status on CD62E+or CD31+/42b- EMPs, or on total microparticles (all P > 0.05). The exploratory analysis revealed that low-fit women had similar changes in EMPs with acute exercise. We concluded that acute moderate-intensity exercise reduces CD62E+and CD31+/42b- EMPs, as well as total microparticles, in healthy midlife women. These effects occurred despite differences in menopausal status and fitness. NEW & NOTEWORTHY This study demonstrates that acute moderate-intensity exercise reduces activated and apoptotic endothelial microparticles in healthy midlife women.

Circulating biological markers in prediction of clinical outcomes in patients with chronic heart failure

The aim of this study was the creation of an evaluation cardiovascular risk score, based on an analysis of circulating biomarkers of CHF. Methods: It was studied prospectively the incidence of fatal and non-fatal cardiovascular events, as well as the frequency of occurrence of death from any cause in a cohort of 194 patients with chronic heart failure during 3 years of observation. Circulating levels of NT-pro brain natriuretic peptide (NT-pro-BNP), galectin-3, high-sensitivity C-reactive protein (hs-CRP), osteoprotegerin, endothelial apoptotic microparticles (EAM) and endothelial progenitor cells (EPC) were measured at baseline. Results: Median follow-up of patients included in the study was 2.76 years (range = 1.8–3.4 years). There were 285 cardiovascular events determined, including 43 deaths and 242 readmissions. Independent predictors of clinical outcomes in patients with CHF were NT-pro-BNP, galectin-3, hs-CRP, osteoprotegerin, CD14 + CD309 + Tie2 + EPA, EAM and EAM/CD14 + CD309 + EPA ratio. Index of cardiovascular risk was calculated by mathematical summation of all ranks of independent predictors, which occurred in the patients included in the study. The findings showed that the average value of the index of cardiovascular risk in patients with CHF was 3.17 units (95 % CI = 1.65–5.10 units). Kaplan–Meier analysis showed that patients with CHF and the magnitude of the risk of less than 4 units have an advantage in survival when compared with patients for whom obtained higher values of ranks cardiovascular risk score. Conclusion: Assessment biomarker risk score of fatal and non-fatal cardiovascular events, constructed on measurement of circulating NT-pro-BNP, galectin-3, hs-CRP, osteoprotegerin, EAM and the ratio of the EAM/CD14 + CD309 + EPA, allowing ample reliably predict the probability survival of patients with CHF, regardless of age, gender, state of the contractile function of the myocardium of the left ventricle and the number of comorbidities.

The Association of Subclinical Hypothyroidism and Pattern of Circulating Endothelial-Derived Microparticles Among Chronic Heart Failure Patients.

Background:Subclinical hypothyroidism (SH) is diagnosed biochemically by the presence of normal serum free thyroxine concentration, in conjunction with an elevated serum thyroid-stimulating hormone level. Recent studies have demonstrated the frequent association between SH and cardiovascular diseases and risk factors.Objectives:To evaluate the impact of SH on patterns of circulating endothelial-derived microparticles, (EMPs) among chronic heart failure (CHF) patientsPatients and Methods:This is a retrospective study involving a cohort of 388 patients with CHF. Fifty-three CHF subjects had SH and 335 patients were free from thyroid dysfunction. Circulating levels of N-terminal-pro brain natriuretic peptide (NT-proBNP), high-sensitivity C-reactive protein (hs-CRP), thyroid-stimulating hormone (TSH), total and free thyroxine (T4), and triiodothyronine (T3), and endothelial apoptotic microparticles (EMPs), were measured at baseline. SH was defined, according to contemporary clinical guidelines, as a biochemical state associated with an elevated serum TSH level of greater 10 μU/L and normal basal free T3 and T4 concentrations.Results:Circulating CD31+/annexin V+ EMPs were higher in patients with SH compared to those without SH. In contrast, activated CD62E+ EMP numbers were not significantly different between both patient cohorts. Using uni (bi) variate and multivariate age- and gender-adjusted regression analysis, we found several predictors that affected the increase of the CD31+/annexin V+ to CD62E+ ratio in the patient study population. The independent impact of TSH per 6.5 μU/L (odds ratio [OR] = 1.23, P = 0.001), SH (OR = 1.22, P = 0.001), NT-proBNP (OR = 1.19, P = 0.001), NYHA class (OR = 1.09, P = 0.001), hs-CRP per 4.50 mg/L (OR = 1.05, P = 0.001), dyslipidemia (OR = 1.06, P = 0.001), serum uric acid per 9.5 mmol/L (OR = 1.04, P = 0.022) on the increase in the CD31+/annexin V+ to CD62E+ ratio, was determined.Conclusions:We believe that the SH state in CHF patients may be associated with the impaired pattern of circulating EMPs, with the predominantly increased number of apoptotic-derived microparticles.

The predictive role of circulating microparticles in patients with chronic heart failure

AimThe study aim was to evaluate whether circulating microparticles with apoptotic or non-apoptotic phenotypes are useful for risk assessment of 3-year cumulative fatal and non-fatal cardiovascular events in CHF patients. MethodsThe incidence of fatal and non-fatal cardiovascular events, as well as the frequency of occurrence of death from any cause in a cohort of 388 patients with CHF during 3years of observation was studied prospectively. Circulating levels of NT-pro brain natriuretic peptide (NT-pro-BNP), high-sensitivity C-reactive protein (hs-CRP), and endothelial apoptotic microparticles (EMPs) were measured at baseline. ResultsMedian follow-up was 2.32years (IQR=1.8–3.1). During follow-up, 110 cardiovascular events (including 43 fatal cases) were determined. Additionally, 74 subjects were hospitalized repetitively due to worsening CHF and also 16 subjects were readmitted in the hospital due to other cardiovascular reasons. In the univariate logistic regression analysis, the main factors independently related with cumulative endpoints were creatinine, fasting glucose, HbA1c, total cholesterol, uric acid, various types of EPMs, NT-pro-BNP, hs-CRP, NYHA class, decreased left ventricular ejection fraction (LVEF) less 45%, and type 2 diabetes mellitus. In multivariate model NYHA class, decreased LVEF (less 45%), NT-pro-BNP, hs-CRP, CD144+/CD31+/annexin V+ EMPs, and CD31+/annexin V+ EMPs remained statistically significant for cumulative endpoint. Adding of CD144+/CD31+/annexin V+ EMCs and CD31+/annexin V+ EMCs to the standard ABC model may improve the relative IDI for cumulative endpoint by 11.4% and 10.5% respectively. ConclusionApoptotic phenotype of circulating microparticles may relate 3-year combined clinical outcomes in CHF patients.

Increased Circulating Endothelial Apoptotic Microparticle to Endothelial Progenitor Cell Ratio Is Associated with Subsequent Decline in Glomerular Filtration Rate in Hypertensive Patients

BackgroundRecent research indicates hypertensive patients with microalbuminuria have decreased endothelial progenitor cells (EPCs) and increased levels of endothelial apoptotic microparticles (EMP). However, whether these changes are related to a subsequent decline in glomerular filtration rate (GFR) remains unclear.Methods and ResultsWe enrolled totally 100 hypertensive out-patients with eGFR ≥30 mL/min/1.73 m2. The mean annual rate of GFR decline (△GFR/y) was −1.49±3.26 mL/min/1.73 m2 per year during the follow-up period (34±6 months). Flow cytometry was used to assess circulating EPC (CD34+/KDR+) and EMP levels (CD31+/annexin V+) in peripheral blood. The △GFR/y was correlated with the EMP to EPC ratio (r = −0.465, p<0.001), microalbuminuria (r = −0.329, p = 0.001), and the Framingham risk score (r = −0.245, p = 0.013). When we divided the patients into 4 groups according to the EMP to EPC ratio, there was an association between the EMP to EPC ratio and the ΔGFR/y (mean ΔGFR/y: 0.08±3.04 vs. −0.50±2.84 vs. −1.25±2.49 vs. −4.42±2.82, p<0.001). Multivariate analysis indicated that increased EMP to EPC ratio is an independent predictor of ΔeGFR/y.ConclusionsAn increased circulating EMP to EPC ratio is associated with subsequent decline in GFR in hypertensive patients, which suggests endothelial damage with reduced vascular repair capacity may contribute to further deterioration of renal function in patients with hypertension.

SAT0212 Circulating Endothelial Microparticles Reflect Microvascular Impairment in Patients with Systemic Sclerosis

BackgroundEndothelial injury plays a key role in the pathogenesis of systemic sclerosis (SSc), but its quantitative assessment has yet to be established clinically. Circulating endothelial microparticles (EMPs) are considered markers...

Post-Dilution High Convective Transport Improves Microinflammation and Endothelial Dysfunction Independently of the Technique

Background/Aims: We examined the effects of different online hemodiafiltration techniques on microinflammation and endothelial damage/repair. Methods: The study was designed as a prospective crossover study. Flow cytometry was used to measure CD14⁺CD16⁺ monocytes, apoptotic endothelial microparticles (EMPs), and endothelial progenitor cells (EPCs). Results: Patients treated with high-flux hemodialysis showed a marked chronic inflammatory state (HF-HD 11 ± 2) versus healthy subjects (HS 3.9 ± 2.3; p < 0.05). High convective transport, independent of the technique used, improves microinflammatory parameters (OL-HDF 7.3 ± 2.1 or MID 6.5 ± 3.4; p < 0.05) and the endothelial damage/repair balance compared to HF-HD (EPCs HF-HD 0.3 ± 0.2), with no differences found between the two modalities (EPCs OL-HDF 0.6 ± 0.1, MID 0.6 ± 0.2; p < 0.05). Conclusion: An increase in convective transport improves the microinflammatory state and the endothelial damage/repair of these patients independently of the technique used.

Circulating Apoptotic Endothelial Cells and Apoptotic Endothelial Microparticles Independently Predict the Presence of Cardiac Allograft Vasculopathy

Maintenance of endothelial homeostasis may prevent the development of cardiac allograft vasculopathy (CAV). This study investigated whether biomarkers related to endothelial injury and endothelial repair discriminate between CAV-negative and CAV-positive heart transplant recipients. CAV is the most important determinant of cardiac allograft survival and a major cause of death after heart transplantation. Fifty-two patients undergoing coronary angiography between 5 and 15 years after heart transplantation were recruited in this study. Flow cytometry was applied to quantify endothelial progenitor cells (EPCs), circulating endothelial cells (CECs), and endothelial microparticles. Cell culture was used for quantification of circulating EPC number and hematopoietic progenitor cell number and for analysis of EPC function. The EPC number and function did not differ between CAV-negative and CAV-positive patients. In univariable models, age, creatinine, steroid dose, granulocyte colony-forming units, apoptotic CECs, and apoptotic endothelial microparticles discriminated between CAV-positive and CAV-negative patients. The logistic regression model containing apoptotic CECs and apoptotic endothelial microparticles as independent predictors provided high discrimination between CAV-positive and CAV-negative patients (C-statistic 0.812; 95% confidence interval: 0.692 to 0.932). In a logistic regression model with age and creatinine as covariates, apoptotic CECs (p = 0.0112) and apoptotic endothelial microparticles (p = 0.0141) were independent predictors (C-statistic 0.855; 95% confidence interval: 0.756 to 0.953). These 2 biomarkers remained independent predictors when steroid dose was introduced in the model. The high discriminative ability of apoptotic CECs and apoptotic endothelial microparticles is a solid foundation for the development of clinical prediction models of CAV.

Different profile of endothelial cell apoptosis in patients with Klinefelter's syndrome.

The aim of this study was to evaluate the effects of testosterone (T) therapy on the sexual function of middle-aged patients with acquired pre-pubertal hypergonadotropic hypogonadism (HrHy) and patients with Klinefelter syndrome (KS). A selected series of 35 middle-aged hypogonadal patients, including those who had recently been observed during andrological counseling (extracted from our database) and had not yet begun hormonal treatment, was recruited for this evaluation. This series of patients included 20 patients with acquired prepubertal HrHy and 15 KS patients who were matched by age and body mass index. All patients underwent an andrological evaluation, which included the administration of the International Index of Erectile Function 5 (IIEF-5) questionnaire and the evaluation of penile echo color Doppler. In addition, by flow cytometry we evaluated the serum levels of apoptotic endothelial microparticles (EMPa) and the vitronectin receptor (VR) at baseline and 6 months after the onset of T therapy. After 6 months of T therapy, patients with HrHy demonstrated mean IIEF-5 scores and a peak systolic velocity that were significantly greater and a mean acceleration time that was significantly lower (p<0.05) than those of patients with KS (p<0.05). In addition, patients with HrHy showed mean EMPa values and VR serum concentrations that were significantly lower than those of patients with KS (p<0.05). KS patients showed significantly improved IIEF-5 scores and Doppler parameters (p<0.05) but not EMPa or VR serum concentrations following treatment. These results indicate that erectile dysfunction in KS can improve with T therapy, although this improvement is more profound in HrHy patients, and these results also suggest that T therapy does not improve the severity of endothelial cell apoptosis in KS patients.

Circulating microparticles as indicators of peripartum cardiomyopathy

Peripartum cardiomyopathy (PPCM) is associated with high mortality and morbidity. Endothelial damage involving cathepsin-D to form a 16 kDa prolactin (PRL) peptide is pathogenetically relevant. Inhibiting PRL peptide with bromocriptine has yielded promising results. We investigated whether microparticles (MPs) can be quantified in serum as markers for diagnosis and treatment effects in PPCM. Patients with PPCM were compared with age-matched healthy post-partum women (PPCTR), healthy pregnant women (PCTR), healthy non-pregnant women (NPCTR), patients with ischaemic cardiomyopathy (ICM), patients with stable coronary artery disease (CAD) and healthy controls (HCTR). Peripartum cardiomyopathy treated with bromocriptine (PPCM-BR) and with PPCM without bromocriptine-treatment as control (PPCM-BRCTR) were compared. Microparticles were determined by flow cytometry. Endothelial MPs (EMPs) were elevated in PPCM compared with PPCTR, PCTR, and NPCTR, each P< 0.001. They were significantly elevated compared with ICM, CAD, and HCTR (P< 0.001). Pregnancy (PCTR) exhibited only slight increases vs. ICM, CAD, NPCTR, and HCTR. The increase in PPCM was due to an increase of activated but not apoptotic EMPs. Platelet-derived microparticles were highly increased in PPCM compared with ICM (P< 0.001) but 9.3 ± 4.4-fold compared with CAD (P< 0.001). In NPCTR (P< 0.001) compared with NPCTR, the increase was 5.9 ± 1.7-fold (P< 0.001). Microparticles generated from monocytes (MMPs) were increased 2.4 ± 1.8-fold in PPCM compared with PCTR (P< 0.001) and 4.8 ± 3.6-fold compared with CAD (P< 0.001), whereas leucocyte MPs (LMPs) were not significantly elevated. Endothelial microparticles were significantly reduced in PPCM treated additionally with bromocriptine compared with PPCM treated only with heart failure therapy (P< 0.001). Microparticle profiles may in long-term distinguish PPCM from normal pregnancy, heart failure, and vascular diseases and might be a diagnostic marker related to the pathomechanism of PPCM.

Dysfunction of the endothelial-platelet pathway in patients with erectile dysfunction before and after daily treatment with tadalafil

Platelet activation results from the exposure of receptors on the surface of the platelet to specific structures on the vessel wall. The aim of this study was to assess the serum concentrations of apoptotic endothelial microparticles (EMPa) and the vitronectin receptor (VR) in patients with erectile dysfunction (ED) before and after treatment with tadalafil. This study included 50 patients with arterial ED. EMPa and VR levels were measured by using flow cytometry. The CD45(neg)-CD144(pos)-annexin V(Pos) events were defined as EMPa, and the CD51(pos)-CD61(pos) events were defined as VR. Patients with ED were evaluated before and after daily treatment with 5 mg tadalafil for 90 days. Patients with arterial ED had serum concentrations of EMPa and VR significantly higher than the control group at baseline. After tadalafil, the serum concentrations of EMPa and VR of the patients with arterial ED were significantly lower than before treatment, but significantly higher than controls. Patients with arterial ED expressed higher levels of both EMPa that are associated with dysfunction of the arterial endothelial pathway and VR-expressing platelet-endothelial elements. Chronic treatment with tadalafil reduced endothelial apoptosis in these patients and also reduced VR expression.

Endothelial apoptosis decrease following tadalafil administration in patients with arterial ED does not last after its discontinuation

Although it is well known that phosphodiesterase V inhibitors, used to treat patients with ED, can improve the endothelial dysfunction in organic vascular forms, few studies have explored the duration of their effect on the endothelium after discontinuation. Therefore, the purpose of this study was to evaluate the serum concentrations of apoptotic endothelial microparticles (EMPa), selected as a marker of endothelial damage, in patients with arterial ED at baseline, during tadalafil administration and 3 and 6 months after its discontinuation. In all, 50 patients with arterial ED were evaluated at baseline and 1 week after administration of tadalafil 5 mg once daily for 90 days. Clinical (International Index of Erectile Function-5 score), instrumental (dynamic penile echo color Doppler) and flow-cytometric (serum EMPa concentrations) analyses were performed before (T0) and 1 week (T1), 3 months (T2) and 6 months (T3) after tadalafil discontinuation. The events CD45(neg)/CD144(pos)/annexinV(pos) were defined as EMPa. At T0, patients with arterial ED had serum EMPa concentration significantly higher than 20 healthy men (controls). At T1, patients with arterial ED showed a serum EMPa concentration significantly lower than T0. The significant difference was maintained, though reduced, at T2 and completely lost at T3. In conclusion, the administration of tadalafil decreased serum EMPa concentration in patients with arterial ED. This positive effect on the endothelial dysfunction disappeared 6 months after tadalafil discontinuation.

Decreased circulating endothelial progenitor cell levels and function in essential hypertensive patients with electrocardiographic left ventricular hypertrophy

The aim of this study was to explore the role of circulating endothelial progenitor cells (EPCs) and endothelial apoptotic microparticles in hypertensive patients with and without electrocardiographic left ventricular hypertrophy (LVH). Flow cytometry was used to assess endothelial cell apoptosis and circulating EPC level by quantification of circulating EPC markers (defined as CD34(+)CD133(+), CD34(+)KDR(+)) and endothelial apoptotic microparticles (defined as CD31(+)/annexin V(+)) in peripheral blood samples. The LVH was defined by ECG with the Cornell voltage criteria. In total, 128 hypertensive patients (83 men and 45 women, aged 59±14 years) were enrolled in this study, in which 107 patients (84%) showed no electrocardiographic evidence of LVH, and 21 patients (16%) fulfilled the LVH criteria by ECG. There were no significant differences in basic characteristics between the two groups, but hypertensive patients with LVH had a higher urine albumin excretion rate than those without LVH (P=0.027). Furthermore, hypertensive patients with LVH were shown to have decreased circulating EPC numbers (all P<0.05) and adhesive function compared with those without LVH (LVH vs. no LVH: 14±6 vs. 30±6 cells per high-power field, P<0.001). Increased numbers of endothelial apoptotic microparticles were noted in hypertensive patients with LVH (4.2±4.9 vs. 2.4±3.4%, P=0.115), although the difference was not significant. This study showed that essential hypertensive patients with electrocardiographic LVH evidence have decreased circulating EPC numbers and adhesive function compared with those without LVH. These findings may explain the pathogenetic processes that link hypertensive LVH and endothelial injury in cardiovascular disease.

Mediterranean diet reduces endothelial damage and improves the regenerative capacity of endothelium

Endothelial dysfunction is a fundamental step in the atherosclerotic disease process. Activation or injury of the endothelium leads to a variety of inflammatory disorders, including the release of microparticles. Endothelial progenitor cells may contribute to the maintenance of the endothelium by replacing injured mature endothelial cells. We studied the influence of dietary fat on the release of endothelial microparticles (EMPs) and endothelial progenitor cells (EPCs) in elderly subjects. Twenty healthy, elderly subjects (10 men and 10 women) consumed 3 diets following a randomized crossover design, each for 4 wk: a saturated fatty acid diet; a low-fat, high-carbohydrate diet; and a Mediterranean diet (MedDiet) enriched in monounsaturated fatty acids. We investigated total microparticles, EMPs from activated endothelial cells (activated EMPs), EMPs from apoptotic endothelial cells (apoptotic EMPs), EPCs, oxidative stress variables, and ischemic reactive hyperemia (IRH). The MedDiet led to lower total microparticle, activated EMP, and apoptotic EMP concentrations and higher EPC numbers than did the other diets (P < 0.001). We detected lower superoxide dismutase activity (P < 0.001), a higher plasma β-carotene concentration (P < 0.001), and lower urinary isoprostane and plasma nitrotyrosine concentrations after consumption of the MedDiet than after consumption of the other 2 diets (P < 0.05). Furthermore, the occurrence of IRH was higher after consumption of the MedDiet than after consumption of the other 2 diets (P < 0.05). Consumption of the MedDiet induces a reduction in endothelial damage and dysfunction, which is associated with an improvement in the regenerative capacity of the endothelium, in comparison with 2 other diets.

Increased circulating CD31+/annexin V+ apoptotic microparticles and decreased circulating endothelial progenitor cell levels in hypertensive patients with microalbuminuria

Microalbuminuria is associated with an increased risk for all-cause and cardiovascular mortality, but the pathophysiologic mechanism underlying the association between urinary albumin excretion and cardiovascular disease remains unclear. Here, we tested the hypothesis that enhanced endothelial apoptotic microparticles and decreased endothelial progenitor cell (EPC) levels might contribute to the pathophysiology of microalbuminuria or macroalbuminuria in cardiovascular disease. Flow cytometry was used to assess endothelial cell apoptosis and circulating EPC levels by quantification of circulating CD31/annexin V apoptotic microparticles and EPC markers (defined as KDRCD133, CD34CD133, CD34KDR) in peripheral blood. In total, 125 patients with hypertension were enrolled in the study, of whom 80 patients (64%) were with normoalbuminuria (albumin excretion rate of <20 microg/min, overnight urine samples), 35 patients (28%) with microalbuminuria (an albumin excretion rate of 20-200 microg/min), and 10 patients (8%) with macroalbuminuria (an albumin excretion rate >200 microg/min). Compared to hypertensive patients with normoalbuminuria, patients with microalbuminuria or macroalbuminuria had significantly more diabetes (P = 0.005), higher systolic blood pressure (P = 0.018), and elevated serum creatinine levels (P < 0.001). Among the three groups, patients with microalbuminuria or macroalbuminuria had significantly increased CD31/annexin V apoptotic microparticles (1.8 +/- 2.2 versus 3.0 +/- 4.3 versus 5.2 +/- 6.2%, P = 0.044) and decreased circulating EPC numbers (P < 0.05). By multivariate analysis, CD31/annexin V apoptotic microparticle level was an independent predictor of urinary albumin excretion rate in hypertensive patients (P < 0.001). Microparticles isolated from hypertensive patients with microalbuminuria or macroalbuminuria attenuated EPC proliferation, migration, and increased H2O2 production, cellular senescence and apoptosis in comparison with those from hypertensive patients with normoalbuminuria. These findings suggest that hypertensive patients with microalbuminuria or macroalbuminuria have increased endothelial apoptotic microparticles and decreased circulating EPC levels, which may contribute to atherosclerotic disease progression and enhanced cardiovascular risk in hypertensive patients with nephropathy.

Effect of Different Dialysis Modalities on Microinflammatory Status and Endothelial Damage

We studied the relationship between microinflammation and endothelial damage in chronic kidney disease (CKD) patients on different dialysis modalities. Four groups of CKD stage 5 patients were studied: 1) 14 nondialysis CKD patients (CKD-NonD); 2) 15 hemodialysis patients (HD); 3) 12 peritoneal dialysis patients with residual renal function >1 ml/min (PD-RRF >1); and 4) 13 peritoneal dialysis patients with residual renal function <or=1 ml/min (PD-RRF <or=1). Ten healthy subjects served as controls. CD14(+)CD16(+) cells and apoptotic endothelial microparticles (EMPs) were measured by flow cytometry. Serum vascular endothelial growth factor (VEGF) was measured by ELISA. CKD-NonD and HD patients had a higher percentage of CD14(+)CD16(+) monocytes than PD groups and controls. CD14(+)CD16(+) was similar in the PD groups, regardless of their RRF, and controls. The four uremic groups displayed a marked increase in apoptotic EMPs and VEGF compared with controls. Apoptotic EMPs and VEGF were significantly higher in HD patients than in CKD-NonD and both PD groups. However, there were no significant differences between CKD-NonD and the two PD groups. There was a correlation between CD14(+)CD16(+) and endothelial damage in CKD-NonD and HD patients, but not in PD and controls. There was an increase in CD14(+)CD16(+) only in CKD-NonD and HD patients. In these patients, there was a relationship between increased CD14(+)CD16(+) and endothelial damage. These results strongly suggest that other factors unrelated to the microinflammatory status mediated by CD14(+)CD16(+) are promoting the endothelial damage in PD, regardless of their RRF.