Nebivolol improves eNOS function and nitric oxide bioavailability in endothelial cells from African Americans

AJH–May 2005–VOL. 18, NO. 5, PART 2

POSTERS: Endothelial Factors (endothelin, nitric oxide, etc.)

nature of the intrarenal and cardiac activation of ET-1/ ETA/ ETB in experimental model of combined diabetes and hypertension. Key Words: Diabetes, Hypertension, Endothelin-1, Rat, Kidney, Heart

P-480 HUMAN ERYTHROPOIETIN-BINDING PROTEIN AND EFFECTS ON ERYTHROPOIETIN-INDUCED HYPERTENSION Mary S. Lee, John S. Lee, Jong Y. Lee. Medicine, Univ. of MN, Minneapolis, MN. Human erythropoietin-binding protein (Epo-bp) and anti-Epo-bp antibodies (aEpo-bp) were purified to test effects on Epo-induced Hypertension. Purified Epo-bp has a strong binding affinity to 125I-Epo, and unlabeled Epo eliminates the binding (p ⬍ 0.0001). Using our new products Epo-bp and aEpo-bp, we examined circadian stage-dependent effects on blood pressure (BP) on Sprague-Dawley rats, which were randomly assigned into control or Rx groups, and compared to the erythropoietin (Epo) Rx group. BPs were measured just before and immediately after the completion of a 4-week course of twice weekly Epo (50U/kg BW), Epo-bp, aEpo-bp or physiological saline injections. Circadian mean BP in Epo Rx vs. control, Epo-bp, aEpo-bp Rx groups were 136.2 ⫾ 2.3 vs. 116.2 ⫾ 1.7, 118.4 ⫾ 2.1 and 116.6⫾2.1 mm Hg, respectively, each p ⬍ 0.0001. Epo Rx increased hematocrit (HT) markedly overall as compared to the control, Epo-bp and aEpo-bp Rx groups (61.6 vs. 42.7, 43.9 and 44.1%, respectively). Epo-bp or aEpo-bp Rx with Epo had almost no effects on Epo-induced HT increase (61.6 in Epo vs. 58.0% or 59.1%, respectively). Thus, both Epo-bp and aEpo-bp maintained similar BP levels as in the control group without reducing HT levels. Key Words: Erythropoiesis, erythropoietin receptor, Epo-bp, affinity purification, ligand binding, blood pressure. Key Words: Blood Pressure, Epo-bp, Erythropoiesis

P-481 NEBIVOLOL IMPROVES ENOS FUNCTION AND NITRIC OXIDE BIOAVAILABILITY IN ENDOTHELIAL CELLS FROM AFRICAN AMERICANS R P Mason, L Kalinowski, R F Jacob, A M Jacoby, T Malinski. Elucida Research, Beverly, MA; Biochemistry Research Lab, Ohio University, Athens, OH; Department of Medicine, Harvard Medical School, Boston, MA. Background: Increased susceptibility of African Americans to cardiovascular disease and its clinical manifestations has been attributed to reduced endothelial function. We tested the ability of nebivolol, a highly selective ␤1-selective agent with direct vasodilating and antioxidant properties, to impact endothelial activity in African Americans. Methods and Results: The effect of nebivolol on endothelial nitric oxide (NO) bioavailability was tested in human umbilical vein endothelial cells (HUVECs) isolated from age-matched African American and White donors with similar CV risk factors, including family history of hypertension and diabetes. Production of NO/O2-/ONOO- was simultaneously recorded using a highly sensitive tandem of electrochemical nanosensors. The results showed significant differences in the kinetics of the initial rates of NO, O2- and ONOO- release between African Americans and Whites: rate of NO release was ⬃5 times slower in African Americans compared with Whites (94 vs. 505 nmol/L/s), whereas the rates of release were faster ⬃2 times for O2- and ⬃4 times for ONOO(22.1 vs. 9.4 nmol/L/s for O2- and 810 vs. 209 nmol/L/s for ONOO-). Pretreatment with 1.0 mmol/L nebivolol restored NO bioavailability in HUVECs from African Americans with concurrent reductions in O2- and ONOO- release, similar to levels recorded from White donor cells. The

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effects of nebivolol were reproducible, dose-dependent and not observed with atenolol, but were by a NAD(P)H-oxidase inhibitor. Conclusions: These data demonstrate that loss of NO bioavailability in African Americans is related to eNOS uncoupling and excessive superoxide generation, a process that can be reversed with nebivolol, independently of ␤1-selective blockade. Key Words: African Americans, Nebivolol, Nitric Oxide

P-482 MEMBRANE LOCATION OF NEBIVOLOL CONTRIBUTES TO ANTIOXIDANT ACTIVITY AND ENDOTHELIAL NITRIC OXIDE RELEASE IN STROKE-PRONE HYPERTENSIVE RATS R P Mason, C Day, R F Jacob, M F Walter, T Malinski. Elucida Research, Beverly, MA; Biochemistry Research Lab, Ohio University, Athens, OH; Department of Medicine, Harvard Medical School, Boston, MA. Background: We tested the ability of nebivolol, a ␤1-selective blocker with direct vasodilating properties, to reverse endothelial dysfunction in Stroke-prone Hypertensive Rat (SHR). The effects of nebivolol on endothelial-dependent nitric oxide (NO) release were correlated with cellular redox state, NO synthase efficiency and molecular membrane localization. Methods and Results: The effect of nebivolol on NO bioavailability, a key indicator of endothelial function, was measured using a porphyrinic nanosensor placed into the lumens of vascular rings from small peripheral arteries. As compared to normal rats, a marked deficiency (⬎40%, p ⬍ 0.01) in NO release was measured in SHR following stimulation with either acetylcholine or calcium ionophore. Nebivolol treatment restored the level of NO release in SHRs to that seen in control animals. In addition, greater NO release was seen with the enantiomer (l-nebivolol) that lacks ␤1-adrenergic activity (340 nM NO versus 180 nM NO for the ␤1-selective d-nebivolol). The mechanism for nebivolol activity may be due to its distinct membrane location that facilitates potent antioxidant scavenging properties, as determined by x-ray diffraction. The antioxidant activity of nebivolol and its enantiomers were not reproduced by other ␤-blockers at low concentrations. Conclusions: Nebivolol is a potent antioxidant that reversed eNOS uncoupling and restored endothelial-dependent NO release in arteries from hypertensive animals prone to stroke, independently of ␤1-blockade activity. Key Words: Antioxidant, Nebivolol, Nitric Oxide

P-483 UTILITY OF A NOVEL RESEARCH CARDIOVASCULAR PROFILING SYSTEM IN ENDOTHELIAL DYSFUNCTION Jose Saban-Ruiz, Begonia Monge, Olivia Sanchez, Rosa Fabregate, Eva Fernandez, David Coca, Arturo Ugalde, Jorge Haurie, Judith Marquez. Endothelial Pathology Unit, Ramon y Cajal Hospital, Madrid, Spain. Introduction: The elasticity of blood vessels may be obtained by a novel non-invasive method using an HDI/PulseWave CR-2000 Research Cardiovascular Profiling System. Not many studies analyze this system as a reliable way of obtaining information about endothelial function. Aims: 1- To determine the prevalence of abnormal large and/or small artery elasticity index in a high cardiovascular risk population. 2- To determine its correlation with macrovascular and/or microvascular endothelial dysfunction. Methods: N⫽94 patients, aged 27-81 yrs (58,83⫹-1,44 SEM), 52,8 % males, 61 hypertensives. Large (C1) and small (C2) artery elasticity index: HDI/PulseWave CR-2000. Macrovascular study: Endothelium (ED) and non-endothelium (NED) dependent vasodilation. Celermajer