Natriuretic peptides and endothelin-1 in patients undergoing coronary artery bypass grafting

Off-pump coronary artery bypass grafting (CABG) is an alternative to conventional CABG using cardiopulmonary bypass. Off-pump technique reduces the complications of CABG performed with extracorporeal circulatory assistance (Lancey et al. 2000; Mack et al. 2004a,b). The object of this study was to compare peri- and postoperative time courses of vasoactive peptides - atrial natriuretic poptide (ANP), brain natriuretic poptide (BNP) and endothelin-1 (ET-1) in off-pump versus on-pump CABG. 22 patients, who underwent on-pump (group A, n = 11) or off-pump CABG (group B, n = 11) were studied. The peri- and postoperative time courses of plasma ANP and BNP were similar in both groups. A statistically significant difference between ET-1 plasma level 2 h after surgery in the group A and ET-1 plasma level 2 h after surgery in the group B (2.46 + or - 1.14 pg/ml/Ht versus 0.74 + or - 0.09 pg/ml/Ht, p < 0.0001) was found. Different CABG techniques were not associated with significant changes in peri- and postoperative plasma ANP and BNP. By contrast, plasma ET-1 significantly rose in the group A 2 h after surgery, indicating endothelial damage.     

Humoral and hemodynamic responses after left ventricular assist device implantation and heart transplantation

Left ventricular assist device (LVAD) implantation and heart transplantation (HTx) are established therapeutic approaches in the treatment of end-stage heart failure. The postoperative humoral responses to the two treatments have not yet been compared. All patients were treated with inhaled nitric oxide (iNO) on weaning from cardiopulmonary bypass as they presented with pulmonary hypertension. We investigated atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), cGMP, endothelin (ET)-1, big endothelin (big ET), and hemodynamic parameters after LVAD implantation (15 patients; age 51 +/- 8 years) or HTx (10 patients; age 53 +/- 6 years) preoperatively, on cardiopulmonary bypass and postoperatively up to 72 hrs after cessation of iNO. Preoperatively, cardiac index (CI), pulmonary artery pressure, pulmonary capillary wedge pressure (PCWP), central venous pressure (CVP), and mean atrial pressure (MAP) were similar for both groups. Similarly, ANP, BNP, cGMP, ET-1, and big ET were comparable before surgery. Seventy-two hours after weaning from iNO, the administered epinephrine dose was higher in the HTx group (P = 0.003); whereas the CVP (P = 0.04) and pulmonary vascular resistance (PVR; P = 0.03) were lower. The following humoral parameters differed markedly: ANP (preoperatively: LVAD, 99 +/- 123 pg/ml; HTx, 197 +/- 199 pg/ml; P = 0.14; vs. 72 hrs after iNO: LVAD, 110 +/- 106 pg/ml; HTx, > 640 +/- 0 pg/ml; P = 0.003) and cGMP (preoperatively: LVAD, 4.4 +/- 5.8 pg/ml; HTx, 5.0 +/- 3.0 pg/ml; P = 0.35; vs. 72 hrs after iNO: LVAD, 8.0 +/- 10.8 pg/ml; HTx, 26.2 +/- 15.8 pg/ml; P = 0.02). Although the hemodynamic effects of both LVAD implantation and HTx in the treatment of end-stage heart failure are comparable, except for the effects on CVP and PVR, the humoral responses with respect to ANP and cGMP were strikingly different. These effects are independent of volume status, iNO, and ETs, suggesting a physiologic response to maintain circulatory homeostasis.     

NT-proBNP and BNP values in cardiac patients with different degree of left ventricular systolic dysfunction

We investigated the performance of brain natriuretic peptides (BNP and NT-proBNP) in detecting various degrees of left ventricular systolic dysfunction. The NT-proBNP assay (Roche) and the BNP assay (Bayer Shionoria) were performed in 46 patients (mean age 50 years; range 20-79 years) with various types of heart disease (chronic heart failure due to coronary artery disease, cardiomyopathy, acquired valve disease, congenital heart diseases) and different impairment of left ventricular systolic dysfunction was assessed by echocardiography. Patients were divided into four groups according to the left ventricular ejection fraction (LVEF) correlated with clinical severity. Significant differences in medians of NT-proBNP and BNP values between all groups were determined (P= 0.0161 for NT-proBNP and P=0.0180 for BNP). For identifying patients with severe systolic dysfunction (LVEF<40%), receiver operating characteristic (ROC) analysis for both BNP and NT-proBNP was performed. The diagnostic performances expressed as areas under the curve were of 0.69 for NT-proBNP (cut off value 367 pg/ml) and 0.60 for BNP (cut off value 172 pg/ml). However, the BNP showed higher sensitivity (85 % vs. 63 %) and a higher positive predictive value (69 % vs 55 %) than the NT-proBNP. The negative predictive values of BNP and NT-proBNP were similar (70 % and 71 % respectively). Brain natriuretic peptides are promising markers for the diagnosis of severe left ventricular systolic dysfunction.     

Endothelin-1, endothelin-1 receptors and cardiac natriuretic peptides in failing human heart

Endothelin (ET)-1 is a potent vasoconstrictor peptide produced in the myocardium that can exert important effects on cardiac myocyte growth and phenotype; cardiac natriuretic peptides (ANP and BNP) are known to act as physiological antagonists of ET-1. In this study a comparative determination of ET-1 receptors and of the local productions of ET-1 and of ANP and BNP was made in different sites of failing and nonfailing hearts. Tissue from right and left atrium, right and left ventricle and interventricular septum from seven adult heart transplant recipients with end-stage idiopathic dilated cardiomyopathy (functional class III and IV, with ejection fraction < 35%) and from four postmortem subjects without cardiac complications was analyzed. In failing hearts we observed a tendency to increase of density of binding sites, most evident in left ventricle (62.6+/-22.6 fmol/mg protein vs. 29.0+/-3.3, mean +/- SEM, p = ns). A prevalence of ET-A subclass, observed in all samples, resulted more pronounced in failing hearts where this increase, found in all the cardiac regions, was more evident in left ventricle (p = 0.0007 vs nonfailing hearts). The local concentrations of ET-1, ANP and BNP resulted significantly increased in failing hearts with respect to controls in all sides of the heart. In failing hearts we have observed a tendency to increase in endothelin receptor density mainly due to a significant upregulation of ET-A subtype and a parallel increase of the tissue levels of ANP, BNP and ET-1 indicating an activation of these systems in heart failure.     

Vascular endothelial growth factor and basic fibroblast growth factor in patients with squamous cell oesophageal cancer

It is suggested that vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) play an important role in tumor-induced angiogenesis. The purpose of this study was to estimate the correlation between VEGF and bFGF levels and tumor pathological status according to pTNM classification in patients with squamous cell oesophageal cancer. A group of 25 healthy controls and 32 consecutive patients with oesophageal cancer were included in this study. Serum VEGF and bFGF levels were determined by enzyme-linked immunosorbent assay (Quantikine R&D Systems). Serum VEGF and bFGF levels were significantly elevated in the patient groups (VEGF: 146.0 pg/ml, 79.0-386.3 pg/ml vs. 38.0 pg/ml, 6.5-135.1 pg/ml, p<0.005, and bFGF: 5.2 pg/ml, 1.2-10.6 pg/ml vs. 2.06 pg/ml, 0.07-4.0 pg/ml, p<0.02 Fisher test). The highest correlation between serum VEGF and bFGF levels were found in patients with advanced cancers, especially with: T4, N1, and M1 factors. The VEGF and bFGF levels were significantly higher in patients with pT4 (p<0.01). Patients with N1 lymph node invasion, compared with N0 factor, have higher levels of angiogenetic factors (p<0.04). Also in patients with advanced cancers with liver metastases the serum levels VEGF and bFGF were significantly higher (M1 vs. M0, VEGF p<0.001 and bFGF p<0.05). Consecutive monitoring of VEGF and bFGF serum levels may be a useful prognostic marker for patients with squamous cell oesophageal cancer.     

Prevalence and misdiagnosis of chronic heart failure in nursing home residents: the role of B-type natriuretic peptides

Background/objectives. Without knowing the exact CHF prevalence, chronic heart failure (CHF) occurs frequently in elderly people both inside and outside nursing homes. For a diagnosis we have to rely on physical examination and additional tests. We therefore run the risk of missing CHF diagnoses or of diagnosing CHF when we should not. Natriuretic peptide assays have emerged as a diagnostic test but their use in nursing home residents is limited. We examined the number of misdiagnoses, the CHF prevalence and the role of natriuretic peptide. Method. Residents in one centre without aphasia, cognitive impairments or metastatic cancer were screened for CHF; the natriuretic peptide levels were measured separately. Results. Of the 150 residents, 103 (64%) were included (79±11 years). The diagnosis of CHF was established in 24 of these 103 residents with NTproBNP 1871 (IQR 539 to 4262) and BNP 194 (IQR 92 to 460) pg/ml. A striking result was that of the 24 residents found to have CHF after the screening, 15 (66%) had previously been undetected: NT-proBNP 1146 (interquartile range (IQR) 228 to 3341) and BNP 200 (IQR 107 to 433) pg/ml. Moreover, in 13 out of 22 residents (62%) who had previously been thought to have CHF, the diagnosis was rejected: NT-proBNP 388 (IQR 174 to 719) and BPN 90 (IQR 35 to 128) pg/ml). Regarding the diagnostic accuracy of NT-proBNP and BNP, the optimal cut-off level of NT-proBNP was 450 pg/ml with a sensitivity of 0.71 and specificity of 0.67, and for BNP it was 100 pg/ml with a sensitivity of 0.71 and specificity of 0.70. Conclusion. Both undetected and incorrect diagnoses of CHF were common. NT-proBNP and BNP were moderately accurate at diagnosing CHF. CHF prevalence was 23%. (Neth Heart J 2008;16: 123-8.)     

Biphasic and monophasic pattern of brain natriuretic peptide release in acute myocardial infarction

This study evaluated brain natriuretic peptide (BNP) release in acute myocardial infarction (AMI), absolute values as well as pattern of its release. There are two different patterns of BNP release in AMI; monophasic pattern--concentration in the first measurement is higher than in the second one, and biphasic pattern--concentration in the first measurement is lower than in the second one. We observed significance of biphasic and monophasic pattern of BNP release related to diagnostic and prognostic value. We included in this prospective observational study total of 75 AMI patients, 52 males and 23 females, average age of 62.3 +/- 10.9 years with range of 42 to 79 years. BNP was measured and pattern of its release was evaluated. In AMI group BNP levels were significantly higher than in controls (462.88 pg/mL vs. 35.36 pg/mL, p < 0.001). We found statistically significant real negative correlation (p < 0.05) between BNP concentration and left ventricle ejection fraction (LVEF) with high correlation coefficient (r = -0.684). BNP concentrations were significantly higher among patients in Killip class II and III compared to Killip class I; Killip class I BNP = 226.18 pg/mL vs. Killip class II 622.51 pg/mL vs. Killip class III 1530.28 pg/mL, p < 0.001. BNP concentrations were significantly higher in patients with; (i) myocardial infarction vs. controls; (BNP 835.80 pg/mL vs. 243.03 pg/mL); (ii) in pts with positive major adverse cardiac events (MACE) vs. negative MACE (BNP 779.08 pg/mL vs. 242.28 pg/mL, p < 0.001); (iii) in pts with positive compared to negative left ventricle (LV) remodelling (BNP 840.77 pg/mL vs. 341.41 pg/mL, p < 0.001). Group with biphasic pattern of BNP release had significantly higher BNP concentration compared to monophasic pattern group. In biphasic pattern group we found significant presence of lower LVEF, Killip class II and III, LV remodelling and MACE. We found that BNP is strong marker of adverse cardiac events in patients presenting with a myocardial infarction. In our AMI group we found significant elevation of BNP and it is suspected that second peak secretion is not only due to systolic dysfunction and subsequent remodeling of LV but also due to impact of ischaemia. Patients with biphasic pattern probably have worse prognosis due to severe coronary heart disease. Besides its diagnostic role as a simple blood marker of systolic function, BNP is also important prognostic marker who helps making clinical decision about early invasive vs. conservative management.     

Relationship of ovarian hormones to hypoxemia in women residents of 4,300 m

Prevalence of excessive erythrocytosis, the main sign of chronic mountain sickness (CMS), is greater in postmenopausal Andean women than in premenopausal women. It is uncertain whether this greater prevalence is related to the decline in female hormones and ventilatory function after the occurrence of the menopause. To study this, we compared the physiological variables involved in the physiopathology of CMS [end-tidal CO(2) (PET(CO(2)), Torr) and end-tidal O(2) (PET(O(2)), Torr), arterial oxygen saturation (Sa(O(2)), %), and Hb concentration (g/dl)] and progesterone and estradiol levels between postmenopausal and premenopausal women, both in the luteal and follicular phases. Women residing in Cerro de Pasco (n = 33; 4,300 m) aged 26--62 yr were studied. Postmenopausal women compared with premenopausal women in the luteal phase had lower PET(O(2)) (48 +/- 4 vs. 53 +/- 2 Torr, P = 0.005) and Sa(O(2)) levels (82 +/- 12 vs. 88 +/- 12%, P < 0.005) and higher PET(CO(2)) (34 +/- 2 vs. 29 +/- 3 Torr, P = 0.005) and Hb concentration (19 +/- 1 vs. 14 +/- 2 g/dl, P < 0.005). In addition, plasma progesterone was negatively correlated with PET(CO(2)) and positively correlated with PET(O(2)) and Sa(O(2)). No clear relationship was found among the cycle phases between estradiol and the variables studied. In conclusion, our results reveal that, before menopause, there is better oxygenation and lower Hb levels in women long residing at altitude, and this is associated with higher levels of progesterone in the luteal phase of the cycle.     

Plasma adrenomedullin and endothelin-1 concentration during low-dose dobutamine infusion: Relationship between pulmonary uptake and pulmonary vascular pressure/flow characteristics

AIM: To study the role of endothelin (ET-1) and adrenomedullin (AM) on pulmonary vascular pressure/flow characteristic (pulmonary arterial pressure/cardiac output (Pap/CO)) during low-dose dobutamine infusion. METHODS: Case control study of 14 patients (12 men, 2 women) with severe lung disease (chronic obstructive pulmonary disease, COPD n=5; cystic fibrosis, CF n=9) and 5 control subjects (CTRL, 4 men, 1 woman). ET-1 and AM plasma levels in pulmonary artery (mixed venous blood, ven) and aorta or femoral artery (arterial, art), were measured at baseline and during dobutamine infusion (5-10-15 mcg kg(-1) min(-1)). The Ppa/CO coordinates obtained at baseline and during dobutamina infusion for each patients were used to calculate the Slope and Intercept by linear regression analysis. RESULTS: Baseline hemodynamics measurements were similar in the three groups with a trend towards a mild elevation in Ppa in CF group (Ppa mm Hg: CTRL 19+/-3.5, COPD 19.4+/-5.5, CF 22.7+/-7.5). Baseline plasma ET-1(ET-1ven pg ml(-1): CTRL 13.9+/-6.7, COPD 20.1+/-14, CF 20.4+/-7.1; ET-1art pg ml(-1): CTRL 16.7+/-6.4, COPD 20.1+/-11.7, CF 18.1+/-3.9) and AM (AMven pg ml(-1): CTRL 15.8+/-5, COPD 31.8+/-17.6, CF 27.7+/-7.6; AMart pg ml(-1): CTRL 15.9+/-1.4, COPD 21.4+/-3.8, CF 27+/-7.6) showed a trend towards higher value among patients' groups compared to the controls. Baseline ET-1 pulmonary gradient did not show significant difference among the three groups as well AM pulmonary gradient. Dobutamine infusion caused a comparable increase of heart rate and CO in the three groups. Mean pulmonary pressure had a trend towards a greater increase in COPD and CF than in controls, consequently, pulmonary Pap/CO relationship showed a steeper slope in patients' groups (Slope mm Hg L(-1) min(-1): CTRL 0.9+/-0.3, COPD 2.1+/-0.8 p<0.02 vs. CTRL, CF 1.9+/-0.9 p<0.03 vs CTRL). During dobutamine plasma ET-1 and AM showed a great individual variability resulting in no significant difference among groups. ET-1 pulmonary gradient showed a trend towards pulmonary uptake in patients' groups (ET-1art-ven pg min(-1): CTRL 2.7+/-2.9, COPD-6.1+/-7.8, CF -4+/-4.8) while AM pulmonary gradient did not show any particular pattern. During dobutamine ET-1 was significantly correlated to Pap/CO characteristics (Slope and ET-1ven, r=-0.59, p<0.05; Slope and ET-1art-ven, r=-0.60, p<0.05; Intercept and ET-1art-ven, r=0.63, p<0.004), and ET-1art-ven was the only independent variable related to Slope and Intercept. CONCLUSIONS: In patients with moderate pulmonary vascular impairment, ET-1 pulmonary gradient, but not AM pulmonary gradient, is inversely correlated with pulmonary incremental resistance, suggesting a role of ET-1 in the regulation of pulmonary vascular resistance.     

Plasma and urinary endothelin-1 concentrations in asphyxiated newborns [corrected

The aim of this study was to determine if there is any correlation between the hypoxia induced deterioration of renal functions and urinary excretions of endothelin (ET). Therefore using a sensitive and specific radioimmunoassay, we have investigated plasma ET-1 concentrations and urine ET-1 excretions in healthy and asphyxiated newborns. Sixteen newborns (10 boys, 6 girls) with perinatal asphyxia or hypoxia of variable seriousness which were followed at Newborn Intensive Care Unit in Eskisehir Osmangazi University Faculty of Medicine were enrolled. Simultaneously, gestation and weight matched 10 newborns (6 boys, 4 girls) with no asphyxia (first minute Apgar score >7) were enrolled as controls. Plasma ET-1 concentrations of the asphyxiated infants (61.8+/-79.3 pg/ml, between 23.4-125.2 pg/ml) were higher than in the control group (29.3+/-22.1 pg/ml, between 12.3 and 50.8 pg/ml, p<0.05). However creatinine clearance values were not different between the two groups (p>0.05), mean fractional excretion of sodium levels (FeNa%) were higher in the study group than the controls (p<0.01). Urinary ET-1 concentrations in the asphyxiated infants were 144.6+/-63.4 pg/ml versus 70.1+/-27.7 pg/ml in the control group (p<0.001). The ET clearance were more elevated in the asphyxiated newborns than in the healthy infants (p<0.05). Urinary ET-1/Cr ratio in the hypoxic infants were significantly elevated in the first day of life when compared with those of healthy infants (p<0.05). Total ET excretion was negatively correlated with FeNa (%) (r=-0.603, p<0.05). Plasma ET-1 concentrations of the asphyxiated infants reduced at 48 hours of age (p<0.001). Fifth minute Apgar score was negatively correlated with urinary ET-1 levels (r=-0.615, p<0.01), urinary Na excretion (r=-0.583, p<0.01), FeNa (%) (r=-0.597, p<0.01) and total ET excretion (r=-0.560, p<0.01) and positively correlated with ET clearance (r=0.559, p<0.05). Urinary ET-1 levels were negatively correlated with umbilical artery BE levels (r=-0.612, p<0.05). To our study, elevated urinary ET-1 levels were observed during perinatal asphyxia and urinary ET-1 levels were negatively correlated with 5th minute Apgar score and cord blood base excess levels. For this reason urinary ET-1 levels could be a marker of perinatal asphyxia as cord blood ET-1 levels. With investigations showing renal production is independent from plasma and increased urinary ET-1/Cr levels in newborn with perinatal asphyxia and also negative correlation between the total ET excretion and FeNa, urinary ET-1 levels could be served as a useful marker to detecting also impaired renal functions in infants with perinatal asphyxia.     

Oxidative stress disrupts nitric oxide synthase activation in liver endothelial cells

Oxidative stress may mediate vascular disruption associated with a loss of endothelial nitric oxide synthase (eNOS) activity and a hypersensitivity to the constrictor effects of endothelin-1 (ET-1). We hypothesize that this is due, in part, to uncoupling of ET(B) receptors from eNOS activation. Thus, we tested whether oxidative stress (OS) affects liver vascular relaxation by reducing basal and ET-1-induced NO production. Primary sinusoidal endothelial cell cultures were pretreated with H(2)O(2) (25 microM) for 1 or 6 h before a 10-min ET-1 stimulation. OS resulted in a significant basal and ET-1-induced decrease in NO production. Acute OS increased the monomeric form of the inhibitory protein caveolin-1 (1.2 +/- 0.05 vs 0.9 +/- 0.02, p < 0.01) and increased the eNOS-caveolin association as determined by coimmunoprecipitation (1.24 +/- 0.04 vs 0.97 +/- 0.04, p < 0.05). ET-1 stimulation further exacerbated these effects. Subacute OS inhibited ET-1-induced eNOS phosphorylation of serine 1177 (activation residue) (1 +/- 0.07 vs 1.6 +/- 0.04, p < 0.05) and dephosphorylation of the inhibitory residue threonine 495 (1.5 +/- 0.08 vs 0.7 +/- 0.02, p < 0.01). Additionally subacute OS resulted in dissociation of eNOS from ET(B) (0.8 +/- 0.09 vs 1.2 +/- 0.06, p < 0.05). Our findings indicate that acute and subacute oxidative stress result in the inhibition of induced nitric oxide synthase activity through distinct mechanisms dependent on caveolin-1 inhibition, ET(B) dissociation, and eNOS phosphorylation.     

Mild hypoxia impairs alveolarization in the endothelial nitric oxide synthase-deficient mouse

In addition to its vasodilator properties, nitric oxide (NO) promotes angiogenesis in the systemic circulation and tumors. However, the role of NO in promoting normal lung vascular growth and its impact on alveolarization during development or in response to perinatal stress is unknown. We hypothesized that NO modulates lung vascular and alveolar growth and that decreased NO production impairs distal lung growth in response to mild hypoxia. Litters of 1-day-old mouse pups from parents that were heterozygous for endothelial nitric oxide synthase (eNOS) deficiency were placed in a hypobaric chamber at a simulated altitude of 12,300 ft (Fi(O(2)) = 0.16). After 10 days, the mice were killed, and lungs were fixed for morphometric and molecular analysis. Compared with wild-type controls, mean linear intercept (MLI), which is inversely proportional to alveolar surface area, was increased in the eNOS-deficient (eNOS -/-) mice [51 +/- 2 micro m (eNOS -/-) vs. 41 +/- 1 micro m (wild type); P < 0.01]. MLI was also increased in the eNOS heterozygote (+/-) mice (44 +/- 1 micro m; P < 0.03 vs. wild type). Vascular volume density was decreased in the eNOS -/- mice compared with wild-type controls (P < 0.03). Lung vascular endothelial growth factor (VEGF) protein and VEGF receptor-1 (VEGFR-1) protein content were not different between the study groups. In contrast, lung VEGFR-2 protein content was decreased from control values by 63 and 34% in the eNOS -/- and eNOS +/- mice, respectively (P < 0.03). We conclude that exposure to mild hypoxia during a critical period of lung development impairs alveolarization and reduces vessel density in the eNOS-deficient mouse. We speculate that NO preserves normal distal lung growth during hypoxic stress, perhaps through preservation of VEGFR-2 signaling.     

Tachycardia-induced myocardial ischemia and diastolic dysfunction potentiate secretion of ANP, not BNP, in hypertrophic cardiomyopathy

The aim of this study was to investigate what factor determines tachycardia-induced secretion of atrial and brain natriuretic peptides (ANP and BNP, respectively) in patients with hypertrophic cardiomyopathy (HCM). HCM patients with normal left ventricular (LV) systolic function and intact coronary artery (n = 22) underwent rapid atrial pacing test. The cardiac secretion of ANP and BNP and the lactate extraction ratio (LER) were evaluated by using blood samples from the coronary sinus and aorta. LV end-diastolic pressure (LVEDP) and the time constant of LV relaxation of tau were measured by a catheter-tip transducer. These parameters were compared with normal controls (n = 8). HCM patients were divided into obstructive (HOCM) and nonobstructive (HNCM) groups. The cardiac secretion of ANP was significantly increased by rapid pacing in HOCM from 384 +/- 101 to 1,268 +/- 334 pg/ml (P < 0.05); however, it was not significant in control and HNCM groups. In contrast, the cardiac secretion of BNP was fairly constant and rather significantly decreased in HCM (P < 0.01). The cardiac ANP secretion was significantly correlated with changes in LER (r = -0.57, P < 0.01) and tau (r = 0.73, P < 0.001) in HCM patients. Tachycardia potentiates the cardiac secretion of ANP, not BNP, in patients with HCM, particularly when it induces myocardial ischemia and LV diastolic dysfunction.     

Plasma endothelin-1 levels and albumin excretion rate in normotensive,microalbuminuric type 2 diabetic patients

BACKGROUND: Endothelin-1 (ET-1) is able to determine functional and structural renal alterations and plasma levels of this vasoconstrictor peptide are increased in diabetic patients. In a selected group of type 2 normotensive diabetic patients with microalbuminuria, we investigated circulating ET-1 levels compared to a control group and verified whether there is a relationship between ET-1 levels and albumin excretion rate in diabetics. SUBJECTS AND METHODS: Thirty-two microalbuminuric type 2 diabetic patients (12 males and 20 females; mean age 57 +/- 8 years) without hypertension, renal failure, hypercholesterolemia or atherosclerotic damage were selected. The control group was made up of 28 healthy subjects matched for sex and age. Blood pressure, creatinine clearance, serum cholesterol and plasma ET-1 values were determined in diabetic and control group. In diabetic patients, glycosilated hemoglobin and urinary albumin excretion rate were also assayed. Mean ET-1 values in diabetics and controls were compared using Student's t-test. Linear regression test was done to relate two variables. Statistical significance was set at p<0.05. RESULTS: Mean ET-1 values were significantly higher in the diabetic group than in controls (11.77 +/- 1.16 pg/ml vs 8.9 +/- 2.1 pg/ml; p<0.05). No relationship (p>0.05) was found between circulating ET-1 and blood pressure, creatinine clearance, serum cholesterol and metabolic control in diabetics. There was a significant positive correlation (r=0.403; p=0.03) between plasma ET-1 levels and albumin excretion rate in diabetic patients. CONCLUSIONS: Our results showed that circulating ET-1 values were increased in microalbuminuric, normotensive, type 2 diabetic patients and correlated with albumin excretion rate. These findings confirm that endothelial dysfunction, as expressed by ET-1 levels, occurs early in these patients and support the hypothesis of a potential role for this peptide in development of microalbuminuria in diabetic nephropathy.     

Markers of Thrombogenesis and Fibrinolysis and Their Relation to Inflammation and Endothelial Activation in Patients with Idiopathic Pulmonary Arterial Hypertension

Background

Chronic anticoagulation is a standard of care in idiopathic pulmonary arterial hypertension (IPAH). However, hemostatic abnormalities in this disease remain poorly understood. Therefore, we aimed to study markers of thrombogenesis and fibrinolysis in patients with IPAH.

Methods

We studied 27 consecutive patients (67% female) with IPAH aged 50.0 years (IQR: 41.0 - 65.0) and 16 controls without pulmonary hypertension. Prothrombin fragment 1+2 (F1+2) and thrombin-antithrombin (TAT) complexes were measured to assess thrombogenesis; tissue-type plasminogen activator (tPA) antigen and plasmin-anti-plasmin complex to characterize activation of fibrinolysis; plasminogen activator inhibitor 1 (PAI-1) to measure inhibition of fibrinolysis; and endothelin-1 (ET-1) and interleukin-6 (IL-6) to assess endothelial activation and systemic inflammation, respectively. In addition, in treatment-naive IPAH patients these markers were assessed after 3 months of PAH-specific therapies.

Results

TPA (10.1[6.8-15.8] vs 5.2[3.3-7.3] ng/ml, p<0.001), plasmin-anti-plasmin (91.5[60.3-94.2] vs 55.8[51.1-64.9] ng/ml, p<0.001), IL-6 (4.9[2.5-7.9] vs 2.1[1.3-3.8] pg/ml, p=0.001) and ET-1 (3.7 [3.3-4.5] vs 3.4[3.1-3.5], p= 0.03) were higher in patients with IPAH than in controls. In IPAH patients plasmin-anti-plasmin and tPA correlated positively with IL-6 (r=0.39, p=0.04 and r=0.63, p<0.001, respectively) and ET-1 (r=0.55, p=0.003 and r=0.59, p=0.001, respectively). No correlation was found between tPA or plasmin-anti-plasmin and markers of thrombogenesis. Plasmin-anti-plasmin decreased after 3 months of PAH specific therapy while the other markers remained unchanged.

Conclusions

In the present study we showed that markers of fibrynolysis were elevated in patients with IPAH however we did not find a clear evidence for increased thrombogenesis in this group of patients. Fibrinolysis, inflammation, and endothelial activation were closely interrelated in IPAH.     

Contributory role of VEGF overexpression in endothelin-1-induced cardiomyocyte hypertrophy

Although endothelin-1 (ET-1) stimulates vascular endothelial growth factor (VEGF) expression in a variety of cells, including endothelial cells and vascular smooth muscle cells, the effects of ET-1 on expression of VEGF and its receptors in cardiomyocytes are unknown. In the present study, we found that treatment of neonatal rat cardiomyocytes with ET-1 for 24 h resulted in upregulation of VEGF and its two principal receptors, fetal liver kinase 1 and fms-like tyrosine kinase 1, in a concentration-dependent manner (10(-12) to 10(-6) M). ET-1 treatment also caused significant cardiomyocyte hypertrophy, as indicated by increases in cell surface area and [(14)C]leucine uptake by cardiomyocytes. Treatment with TA-0201 (10(-6) M), an ET(A)-selective blocker, eliminated ET-1-induced overexpression of VEGF and its receptors as well as cardiomyocyte hypertrophy. Treatment with VEGF neutralizing peptides (5-10 mug/ml) partially but significantly inhibited ET-1-induced cardiomyocyte hypertrophy. These results suggest that ET-1 treatment of cardiomyocytes promotes overexpression of VEGF and its receptors via activation of ET(A) receptors, and consequently the upregulated VEGF signaling system appears to contribute, at least in part, to ET-1-induced cardiomyocyte hypertrophy.     

Endothelin-1 mediates cardiac mast cell degranulation, matrix metalloproteinase activation, and myocardial remodeling in rats

The objective of this study was to determine whether elevated circulating levels of endothelin (ET)-1 are capable of mediating left ventricular (LV) mast cell degranulation and thereby induce matrix metalloproteinase (MMP) activation. After the administration of 20 pg/ml ET-1 to blood-perfused isolated rat hearts, LV tissue was analyzed for signs of mast cell degranulation and MMP activation. Relative to control, ET-1 produced extensive mast cell degranulation as well as a significant increase in myocardial water content (78.8 +/- 1.5% vs. 74.2 +/- 2.2%, P <0.01), a marked 107% increase in MMP-2 activity (P <0.05), and a substantial decrease in collagen volume fraction (0.69 +/- 0.09% vs. 0.99 +/- 0.04%, P <0.001). Although the myocardial edema would be expected to increase ventricular stiffness, compliance was not altered, and moderate ventricular dilatation was observed (end-diastolic volume at end-diastolic pressure of 0 mmHg of 330.2 +/- 22.1 vs. 298.9 +/- 17.4 microl in ET-1 treated vs. control, respectively, P=0.07). Additionally, pretreatment with the mast cell stabilizer nedocromil prevented ET-1-induced changes in MMP-2 activity, myocardial water content, collagen volume fraction, and end-diastolic volume. These findings demonstrate that ET-1 is a potent cardiac mast cell secretogogue and further indicate that ET-1-mediated mast cell degranulation is a potential mechanism responsible for myocardial remodeling.     

Passive leg movement enhances interstitial VEGF protein, endothelial cell proliferation, and eNOS mRNA content in human skeletal muscle

The present study used passive limb movement as an experimental model to study the effect of increased blood flow and passive stretch, without enhanced metabolic demand, in young healthy male subjects. The model used was 90 min of passive movement of the leg leading to a 2.8-fold increase (P < 0.05) in blood flow without a significant enhancement in oxygen uptake. Muscle interstitial fluid was sampled with microdialysis technique and analyzed for vascular endothelial growth factor (VEGF) protein and for the effect on endothelial cell proliferation. Biopsies obtained from the musculus vastus lateralis were analyzed for mRNA content of VEGF, endothelial nitric oxide synthase (eNOS), and matrix metalloproteinase-2 (MMP-2). The passive leg movement caused an increase (P < 0.05) in interstitial VEGF protein concentration above rest (73 +/- 21 vs. 344 +/- 83 pg/ml). Addition of muscle dialysate to cultured endothelial cells revealed that dialysate obtained during leg movement induced a 3.2-fold higher proliferation rate (P < 0.05) than dialysate obtained at rest. Passive movement also enhanced (P < 0.05) the eNOS mRNA level fourfold above resting levels. VEGF mRNA and MMP-2 mRNA levels were unaffected. The results show that a session of passive leg movement, elevating blood flow and causing passive stretch, augments the interstitial concentrations of VEGF, the proliferative effect of interstitial fluid, and eNOS mRNA content in muscle tissue. We propose that enhanced blood flow and passive stretch are positive physiological stimulators of factors associated with capillary growth in human muscle.     

Assessment of pro- and antiangiogenic factors blood serum concentrations in patients with hormonal inactive adrenal tumors

INTRODUCTION: The growth and persistence of solid tumors and their metastases is connected with angiogenesis. This process is determined by activity of pro- and antyangiogenic factors. VEGF is the one of the most important factors having a stimulant effect on angiogenesis. Soluble forms of VEGF receptors are inhibitors of angiogenesis. The soluble forms of VEGF receptors containing extra cellular part of receptor, which binds ligand, seem to be real inhibitors of VEGF. THE AIM OF THE STUDY: Evaluation the value of serum VEGF and soluble forms of VEGF receptors concentration as a marker of malignancy in patients with hormonal inactive adrenal tumors. MATERIAL AND METHODS: Twenty seven patients (18 female, 9 male; mean age 48+/-4.3 years) with adrenocortical carcinoma (N=8), adrenal metastases (N=4) and adrenocortical adenoma (N=15) were included in this study. Age- and gender-matched control samples were acquired from healthy volunteers (N=10). Serum VEGF and sVEGFR levels were determinated by means of ELISA assay. Statistical analysis was performed using the Student-t test, the Pearson's test and the series test. RESULTS: In healthy controls mean VEGF level was 197.2 pg/ml, sVEGFR-1 43.5 pg/ml and sVEGFR-2 8976.3 pg/ml. Patients with adrenocortical carcinoma had the levels of VEGF (1263.8 pg/ml) significantly higher and of sVEGFR-2 (5893.7 pg/ml) significantly lower in comparison to control group (p<0.05). On the other hand the mean VEGF (334.2 pg/ml) concentration in patients with benign adrenocortical adenoma wasn't significant different than in control group (p>0.05) but mean sVEGFR-1 (21.7 pg/ml) and sVEGFR-2 (7106.4 pg/ml) concentrations were significantly lower than in the control (p<0.05). In metastases group mean VEGF (485.9 pg/ml) level was higher and sVEGFR-2 (5455.2 pg/ml) was lower than in control group (p<0.05). CONCLUSION: These data suggest that determination of VEGF and sVEGFR concentration in the serum of patients with hormonal inactive adrenal tumors may be applied as an additional marker of malignancy.