Showing papers by "Edward G. Lakatta published in 2006"

Resveratrol improves health and survival of mice on a high-calorie diet

Abstract: Resveratrol (3,5,49-trihydroxystilbene) extends the lifespan of diverse species including Saccharomyces cerevisiae, Caenorhabditis elegans and Drosophila melanogaster. In these organisms, lifespan extension is dependent on Sir2, a conserved deacetylase proposed to underlie the beneficial effects of caloric restriction. Here we show that resveratrol shifts the physiology of middle-aged mice on a high-calorie diet towards that of mice on a standard diet and significantly increases their survival. Resveratrol produces changes associated with longer lifespan, including increased insulin sensitivity, reduced insulin-like growth factor-1 (IGF-I) levels, increased AMP-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor-c coactivator 1a (PGC-1a) activity, increased mitochondrial number, and improved motor function. Parametric analysis of gene set enrichment revealed that resveratrol opposed the effects of the high-calorie diet in 144 out of 153 significantly altered pathways. These data show that improving general health in mammals using small molecules is an attainable goal, and point to new approaches for treating obesity-related disorders and diseases of ageing.

Central Role for the Cardiotonic Steroid Marinobufagenin in the Pathogenesis of Experimental Uremic Cardiomyopathy

Abstract: Patients with chronic renal failure develop a "uremic" cardiomyopathy characterized by diastolic dysfunction, cardiac hypertrophy, and systemic oxidant stress. Patients with chronic renal failure are also known to have increases in the circulating concentrations of the cardiotonic steroid marinobufagenin (MBG). On this background, we hypothesized that elevations in circulating MBG may be involved in the cardiomyopathy. First, we observed that administration of MBG (10 microg/kg per day) for 4 weeks caused comparable increases in plasma MBG as partial nephrectomy at 4 weeks. MBG infusion caused increases in conscious blood pressure, cardiac weight, and the time constant for left ventricular relaxation similar to partial nephrectomy. Decreases in the expression of the cardiac sarcoplasmic reticulum ATPase, cardiac fibrosis, and systemic oxidant stress were observed with both MBG infusion and partial nephrectomy. Next, rats were actively immunized against a MBG-BSA conjugate or BSA control, and partial nephrectomy was subsequently performed. Immunization against MBG attenuated the cardiac hypertrophy, impairment of diastolic function, cardiac fibrosis, and systemic oxidant stress seen with partial nephrectomy without a significant effect on conscious blood pressure. These data suggest that the increased concentrations of MBG are important in the cardiac disease and oxidant stress state seen with renal failure.

High Basal Protein Kinase A–Dependent Phosphorylation Drives Rhythmic Internal Ca2+ Store Oscillations and Spontaneous Beating of Cardiac Pacemaker Cells

Abstract: Local, rhythmic, subsarcolemmal Ca2+ releases (LCRs) from the sarcoplasmic reticulum (SR) during diastolic depolarization in sinoatrial nodal cells (SANC) occur even in the basal state and activate an inward Na(+)-Ca2+ exchanger current that affects spontaneous beating. Why SANC can generate spontaneous LCRs under basal conditions, whereas ventricular cells cannot, has not previously been explained. Here we show that a high basal cAMP level of isolated rabbit SANC and its attendant increase in protein kinase A (PKA)-dependent phosphorylation are obligatory for the occurrence of spontaneous, basal LCRs and for spontaneous beating. Gradations in basal PKA activity, indexed by gradations in phospholamban phosphorylation effected by a specific PKA inhibitory peptide were highly correlated with concomitant gradations in LCR spatiotemporal synchronization and phase, as well as beating rate. Higher levels of basal PKA inhibition abolish LCRs and spontaneous beating ceases. Stimulation of beta-adrenergic receptors extends the range of PKA-dependent control of LCRs and beating rate beyond that in the basal state. The link between SR Ca2+ cycling and beating rate is also present in vivo, as the regulation of beating rate by local beta-adrenergic receptor stimulation of the sinoatrial node in intact dogs is markedly blunted when SR Ca2+ cycling is disrupted by ryanodine. Thus, PKA-dependent phosphorylation of proteins that regulate cell Ca2+ balance and spontaneous SR Ca2+ cycling, ie, phospholamban and L-type Ca2+ channels (and likely others not measured in this study), controls the phase and size of LCRs and the resultant Na(+)-Ca2+ exchanger current and is crucial for both basal and reserve cardiac pacemaker function.

Matrix Metalloproteinase 2 Activation of Transforming Growth Factor-β1 (TGF-β1) and TGF-β1–Type II Receptor Signaling Within the Aged Arterial Wall

Abstract: Objective— To study matrix metalloproteinase 2 (MMP-2) effects on transforming growth factor-β1 (TGF-β1) activation status and downstream signaling during arterial aging. Methods and Results— Western blotting and immunostaining showed that latent and activated TGF-β1 are markedly increased within the aorta of aged Fisher 344 cross-bred Brown Norway (30 months of age) rats compared with adult (8 months of age) rats. Aortic TGF-β1–type II receptor (TβRII), its downstream molecules p-similar to mad-mother against decapentaplegic (SMAD)2/3 and SMAD4, fibronectin, and collagen also increased with age. Moreover, TGF-β1 staining is colocalized with that of activated MMP-2 within the aged arterial wall and vascular smooth muscle cell (VSMC) in vitro, and this physical association was confirmed by coimmunoprecipitation. Incubation of young aortic rings ex vivo or VSMCs in vitro with activated MMP-2 enhanced active TGF-β1, collagen, and fibronectin expression to the level of untreated old counterparts, and this effect was abolished via inhibitors of MMP-2. Interestingly, in old untreated rings or VSMCs, the increased TGF-β1, fibronectin, and collagen were also substantially reduced by inhibition of MMP-2. Conclusions— Active TGF-β1, its receptor, and receptor-mediated signaling increase within the aortic wall with aging. TGF-β1 activation is dependent, in part at least, by a concomitant age-associated increase in MMP-2 activity. Thus, MMP-2–activated TGF-β1, and subsequently TβRII signaling, is a novel molecular mechanism for arterial aging.

Relationship between androgenic hormones and arterial stiffness, based on longitudinal hormone measurements

Abstract: Circulating testosterone levels (T) decrease with age in men. Low T has been associated with coronary disease and with risk factors for atherosclerosis. This study examines the relationship in men ...

Membrane Potential Fluctuations Resulting From Submembrane Ca2+ Releases in Rabbit Sinoatrial Nodal Cells Impart an Exponential Phase to the Late Diastolic Depolarization That Controls Their Chronotropic State

Abstract: Stochastic but roughly periodic LCRs (Local subsarcolemmal ryanodine receptor–mediated Ca 2+ Releases) during the late phase of diastolic depolarization (DD) in rabbit sinoatrial nodal pacemaker cells (SANCs) generate an inward current ( I NCX ) via the Na + /Ca 2+ exchanger. Although LCR characteristics have been correlated with spontaneous beating, the specific link between LCR characteristics and SANC spontaneous beating rate, ie, impact of LCRs on the fine structure of the DD, have not been explicitly defined. Here we determined how LCRs and resultant I NCX impact on the DD fine structure to control the spontaneous SANC firing rate. Membrane potential ( V m ) recordings combined with confocal Ca 2+ measurements showed that LCRs impart a nonlinear, exponentially rising phase to the DD later part, which exhibited beat-to-beat V m fluctuations with an amplitude of approximately 2 mV. Maneuvers that altered LCR timing or amplitude of the nonlinear DD (ryanodine, BAPTA, nifedipine or isoproterenol) produced corresponding changes in V m fluctuations during the nonlinear DD component, and the V m fluctuation response evoked by these maneuvers was tightly correlated with the concurrent changes in spontaneous beating rate induced by these perturbations. Numerical modeling, using measured LCR characteristics under these perturbations, predicted a family of local I NCX that reproduced V m fluctuations measured experimentally and determined the onset and amplitude of the nonlinear DD component and the beating rate. Thus, beat-to-beat V m fluctuations during late DD phase reflect the underlying LCR/ I NCX events, and the ensemble of these events forms the nonlinear DD component that ultimately controls the SANC chronotropic state in tight cooperation with surface membrane ion channels.

Erythropoietin, Modified to Not Stimulate Red Blood Cell Production, Retains Its Cardioprotective Properties

Abstract: Erythropoietin (EPO), a hematopoietic cytokine, possesses strong antiapoptotic, tissue-protective properties For clinical applications, it is desirable to separate the hematopoietic and tissue-protective properties Recently introduced carbamylated erythropoietin (CEPO) does not stimulate the erythropoiesis but retains the antiapoptotic and neuroprotective effects We tested the ability of CEPO to protect cardiac tissue from toxin-induced and oxidative stress in vitro and ischemic damage in vivo and compared these effects with the effects of EPO CEPO reduced by 50% the extent of staurosporine-induced apoptosis in isolated rats' cardiomyocytes and increased by 25% the reactive oxygen species threshold for induction of the mitochondrial permeability transition In an experimental model of myocardial infarction induced by permanent ligation of a coronary artery in rats, similarly to EPO, a single bolus injection of 30 mug/kg bwt of CEPO immediately after coronary ligation reduced apoptosis in the myocardial area at risk, examined 24 h later, by 50% Left ventricular remodeling (ventricular dilation) and functional decline (fall in ejection fraction) assessed by repeated echocardiography were significantly and similarly attenuated in CEPO- and EPO-treated rats Four weeks after coronary ligation, the myocardial infarction (MI) size in CEPO- and EPO-treated rats was half of that in untreated coronary-ligated animals Unlike EPO, CEPO had no effect on hematocrit The antiapoptotic cardioprotective effects of CEPO, shown by its ability to limit both post-MI left ventricular remodeling and the extent of the myocardial scar in the model of permanent coronary artery ligation in rats, demonstrate comparable potency to that of native (nonmodified) EPO

The integration of spontaneous intracellular Ca2+ cycling and surface membrane ion channel activation entrains normal automaticity in cells of the heart's pacemaker.

Abstract: Although the ensemble of voltage- and time-dependent rhythms of surface membrane ion channels, the membrane "Clock", is the immediate cause of a sinoatrial nodal cell (SANC) action potential (AP), it does not necessarily follow that this ion channel ensemble is the formal cause of spontaneous, rhythmic APs. SANC also generates intracellular oscillatory spontaneous Ca 2+ releases that ignite excitation (SCaRIE) of the surface membrane via Na + /Ca 2+ exchanger activation. The idea that a rhythmic intracellular Ca 2+ Clock might keep time for normal automaticity of SANC, however, has not been assimilated into mainstream pacemaker dogma. Recent experimental evidence, derived from simultaneous, confocal imaging of submembrane Ca 2+ and membrane potential of SANC, and supported by numerical modeling, indicates that normal automaticity of SANC is entrained and stabilized by the tight integration of the SR Ca 2+ Clock that generates rhythmic SCaRIE, and the surface membrane Clock that responds to SCaRIE to immediately produce APs of an adequate shape. Thus, tightly controlled, rhythmic SCaRIE does not merely fine tune SANC AP firing, but is the formal cause of the basal and reserve rhythms, insuring pacemaker stability by rhythmically integrating multiple Ca 2+ -dependent functions, and effects normal automaticity by rhythmic ignition of the surface membrane Clock.

Endogenous Na/K-ATPase inhibitors in patients with preeclampsia.

Abstract: Although preeclampsia (PE) is a major cause of maternal and fetal mortality, its pathogenesis is not fully understood. Digitalis-like cardiotonic steroids (CTS) are believed to be involved in the pathophysiology of PE, as illustrated by clinical observations that DIGIBIND, a digoxin antibody which binds CTS, lowers blood pressure in PE. Recently we reported that plasma levels of marinobufagenin (MBG), a vasoconstrictor CTS, are increased fourfold in patients with severe PE. In the present study, we tested whether anti-MBG, or anti-ouabain antibodies, or DIGIBIND can reverse inhibition of erythrocyte Na/K-ATPase (NKA) from patients with mild PE (blood pressure, 149 +/- 3/93 +/- 3 mm Hg; age, 28 +/- 2 years; gestational age, 37 +/- 1 weeks). Development of PE was associated with twofold rise in plasma MBG levels (1.58 +/- 0.15 vs. 0.80 +/- 0.11 nmol/L; P 0.5). The present observations provide evidence for a role for MBG in the pathogenesis of PE, and suggest that antibodies against MBG may be useful in the treatment of this syndrome.

ANP Differentially Modulates Marinobufagenin-Induced Sodium Pump Inhibition in Kidney and Aorta

Abstract: NaCl loading and plasma volume expansion stimulate 2 natriuretic systems, vasoconstrictor, digitalis-like Na/K-ATPase inhibitors and vasorelaxant ANP peptides. Several hormones, including ANP, regulate activity of the Na/K-ATPase by modulation of its phosphorylation state. We studied effects of ANP on Na/K-ATPase phosphorylation and inhibition by an endogenous sodium pump ligand, marinobufagenin, in the aorta and renal medulla from male Sprague-Dawley rats. Marinobufagenin dose-dependently inhibited the Na/K-ATPase in renal and vascular membranes at the level of higher (nanomolar) and lower affinity (micromolar) binding sites. Marinobufagenin (1 nmol/L) inhibited Na/K-ATPase in aortic sarcolemma (18%) and in renal medulla (19%). prepro-ANP 104 to 123 (ppANP) and -human ANP ((-hANP) both 1 nmol/L) potentiated marinobufagenin-induced Na/K-ATPase inhibition in the kidney, but reversed the effect of marinobufagenin in the aorta. Similarly, ppANP and -hANP modulated the sodium pump (ouabain-sensitive 86 Rb uptake) inhibitory effects of marinobufagenin in the aorta and renal medulla. In renal medulla, ppANP and -hANP induced -1 Na/K-ATPase phosphorylation, whereas in aorta, both peptides dephosphorylated Na/K-ATPase. The effect of ppANP on Na/K-ATPase phosphorylation and inhibition was mimicked by a protein kinase G activator, 8-Br-PET-cGMP (10 mol/L), and prevented by a protein kinase G inhibitor, KT5823 (60 nmol/L). Our results suggest that -1 Na/K-ATPase inhibition by marinobufagenin in the kidney is enhanced via Na/K-ATPase phosphorylation by ANP, whereas in the aorta, ANP exerts an opposite effect. The concurrent production of a vasorelaxant, ANP, and a vasoconstrictor, marinobufagenin, potentiate each other's natriuretic effects, but ANP peptides may offset the deleterious vasoconstrictor effect of marinobufagenin. (Hypertension. 2006;48:1160-1168.)

Uremic Cardiomyopathy—An Endogenous Digitalis Intoxication?: Central Role for the Cardiotonic Steroid Marinobufagenin in the Pathogenesis of Experimental Uremic Cardiomyopathy. Hypertension 47: 488–495, 2006

Abstract: Since the days of Richard Bright ([1][1]), it has been common knowledge that the heart is a target organ in uremia, and by the beginning of the 20th century it had been recognized that heart failure and cardiac death were frequent in renal patients ([2][2]). Despite the common occurrence of failing

Therapeutic Effectiveness of a Single vs Multiple Doses of Erythropoietin After Experimental Myocardial Infarction in Rats

Abstract: Background Systemic application of recombinant human erythropoietin (rhEPO) greatly limits cardiac tissue damage and attenuates left ventricular (LV) remodeling after experimentally induced myocardial infarction (MI). However, multiple injections of rhEPO stimulate red blood cell production and elevate the hematocrit (Htc), which might negatively affect the outcome of acute MI. We compared the outcome of experimental MI in rats treated with a single or multiple doses of rhEPO.

Anti-marinobufagenin antibodies and methods for their use

Abstract: Described herein are deposited hybridoma cell lines and the monoclonal antibodies produced by these hybridomas, and antigen binding fragments thereof. These monoclonal antibodies and antigen binding fragments specifically bind marinobufagenin. The disclosure also encompasses the use of these monoclonal antibodies or antigen binding fragments in a method for detecting the presence of marinobufagenin in a biological sample. Also provided are methods for the use of these monoclonal antibodies or antigen binding fragments as prophylactic, therapeutic, and diagnostic agents for the detection, inhibition and treatment of a cardiovascular disease, for example, essential hypertension, hypertension associated with preeclampsia, eclampsia, or renal failure, or myocardial fibrosis in a subject.

Calorie restriction reduces MMP-2 activity and retards age-associated aortic restructuring in rats

Abstract: Calorie restriction (CR) is a reproducible approach to extend longevity in rodents. Arterial intimal thickening, with increased matrix-metalloproteinase-type-II (MMP2) activation and changes in other variables in table, that accompany aging, are risk factors for atherosclerotic events, which negatively impact lifespan. However, the effects of CR on arterial restructuring and MMP2 activation with aging remain elusive. To address this question we used the aorta and early passage vascular smooth muscle cells (VSMC) isolated from ad libitum (AL)-fed young (6 mo) and old (24 mo) and age-matched, 40% restricted CR F344 rats. Immunostaining showed that intimal VSMC number was increased by 4.4 fold in old compared to young AL rats (table), but was substantially reduced in the CR rat with aging. The intima-medial collagen deposition was increased by 1.5 fold, and the elastin fraction was decreased by 1.8 fold in the old AL rat. Impressively, aortic collagen and elastin fibers did not significantly change in the CR rat during aging. Notably, age enhanced in situ MMP2 activity within the aortic wall in the AL rat by 6.0 fold, but this was blunted in the aorta of the old CR rats. Additionally, a potent pro-fibrogenic cytokine TGF-s1, a product of MMP-2 cleavage, and its downstream signaling molecule p-SMAD-2 were enhanced by 5.0 fold, and by 3.3 fold respectively, in old compared to young AL rats, but CR reduced this effect. The intima-media gradient of a potent chemoattractant, platelet derived growth factor (PDGF) was increased 2.2 fold in old compared to young AL rats, but this was attenuated in CR rats. In addition, CR decreased early passage VSMC invasive capability in vitro in response to PDGF, both in young (26%) and old (15%). In summary CR retards age-associated arterial restructuring in rats, at least in part, via reduction of MMP2 and TGF-s1 activation, the intima-media PDGF gradient, and VSMC invasive capability.

Abstract 1717: Calorie Restriction Reduces MMP-2 Activity and Retards Age-associated Aortic Restructuring in Rats

Abstract: Calorie restriction (CR) is a reproducible approach to extend longevity in rodents Arterial intimal thickening, with increased matrix-metalloproteinase-type-II (MMP2) activation and changes in oth

ANP differentially modulates marinobufagenin-induced sodium pump inhibition in kidney and aorta. Commentary

Abstract: NaCl loading and plasma volume expansion stimulate 2 natriuretic systems, vasoconstrictor, digitalis-like Na/K-ATPase inhibitors and vasorelaxant ANP peptides. Several hormones, including ANP, regulate activity of the Na/K-ATPase by modulation of its phosphorylation state. We studied effects of ANP on Na/K-ATPase phosphorylation and inhibition by an endogenous sodium pump ligand, marinobufagenin, in the aorta and renal medulla from male Sprague-Dawley rats. Marinobufagenin dose-dependently inhibited the Na/K-ATPase in renal and vascular membranes at the level of higher (nanomolar) and lower affinity (micromolar) binding sites. Marinobufagenin (1 nmol/L) inhibited Na/K-ATPase in aortic sarcolemma (18%) and in renal medulla (19%). prepro-ANP 104 to 123 (ppANP) and α-human ANP ([a-hANP] both 1 nmol/L) potentiated marinobufagenin-induced Na/K-ATPase inhibition in the kidney, but reversed the effect of marinobufagenin in the aorta. Similarly, ppANP and α-hANP modulated the sodium pump (ouabain-sensitive 86 Rb uptake) inhibitory effects of marinobufagenin in the aorta and renal medulla. In renal medulla, ppANP and α-hANP induced α-1 Na/K-ATPase phosphorylation, whereas in aorta, both peptides dephosphorylated Na/K-ATPase. The effect of ppANP on Na/K-ATPase phosphorylation and inhibition was mimicked by a protein kinase G activator, 8-Br-PET-cGMP (10 μmol/L), and prevented by a protein kinase G inhibitor, KT5823 (60 nmol/L). Our results suggest that α-1 Na/K-ATPase inhibition by marinobufagenin in the kidney is enhanced via Na/K-ATPase phosphorylation by ANP, whereas in the aorta, ANP exerts an opposite effect. The concurrent production of a vasorelaxant, ANP, and a vasoconstrictor, marinobufagenin, potentiate each other's natriuretic effects, but ANP peptides may offset the deleterious vasoconstrictor effect of marinobufagenin.