Showing papers by "Edward G. Lakatta published in 1995"

Impact of age on the cardiovascular response to dynamic upright exercise in healthy men and women.

Abstract: To examine whether age differentially modifies the physiological response to exercise in men and women, we performed gated radionuclide ventriculography with measurement of left ventricular volumes...

Functional coupling of the beta 2-adrenoceptor to a pertussis toxin-sensitive G protein in cardiac myocytes.

Abstract: Recently we demonstrated that the effects of beta 2-adrenoceptor (AR) stimulation to augment Ca2+ current (ICa), cytosolic Ca2+ (Cai) transients, and contractility in rat ventricular myocytes are largely dissociated from its effect to increase cellular cAMP levels. This result suggested that beta 2ARs might be coupled to signaling pathways other than the Gs alpha-mediated activation of adenylyl cyclase. Here we show that pertussis toxin (PTX) pretreatment specifically potentiates the responses of rat heart cells to beta 2AR but not beta 1AR stimulation. After PTX pretreatment, 1) the dose-response curve for the effects of the beta 2AR agonist zinterol on contraction amplitude is shifted leftward and upward (EC50 changed from about 1.0 microM to 70 nM), 2) in indo-1-loaded cells, the maximal effects of zinterol (10(-5) M) on Cai transient and contraction amplitudes are additionally increased 1.7- and 2.0-fold, respectively, over those in control cells, and 3) the increase in ICa amplitude induced by the same zinterol concentration is potentiated by 2.5-fold. Similar effects of PTX are observed when beta 2ARs are stimulated by isoproterenol in the presence of a selective beta 1AR blocker, CGP 20712A. All effects of beta 2AR agonists in both PTX-treated and control cells are abolished by a selective beta 2AR blocker, ICI 118,551. In contrast, neither the base-line ICa, Cai transient, and contraction in the absence of beta AR stimulation nor the beta 1AR-mediated augmentations of these parameters are significantly altered by PTX treatment. These results demonstrate, for the first time, that the Gs-coupled beta 2AR can simultaneously activate a pathway that leads to functional inhibition in cardiac cells via a PTX-sensitive G protein. The activation of more than one G protein during beta 2AR stimulation, leading to functionally opposite effects, may provide a mechanism to protect the heart from Ca2+ overload and arrhythmias during the response to stress.

Taxol inhibits neointimal smooth muscle cell accumulation after angioplasty in the rat.

Abstract: Despite significant improvements in the primary success rate of the medical and surgical treatments for atherosclerotic disease, including angioplasty, bypass grafting, and endarterectomy, secondary failure due to late restenosis continues to occur in 30-50% of individuals. Restenosis and the later stages in atherosclerotic lesions are due to a complex series of fibroproliferative responses to vascular injury involving potent growth-regulatory molecules (such as platelet-derived growth factor and basic fibroblast growth factor) and resulting in vascular smooth muscle cell (VSMC) proliferation, migration, and neointimal accumulation. We show here, based on experiments with both taxol and deuterium oxide, that microtubules are necessary for VSMCs to undergo the multiple transformations contributing to the development of the neointimal fibroproliferative lesion. Taxol was found to interfere both with platelet-derived growth factor-stimulated VSMC migration and with VSMC migration and with VSMC proliferation, at nanomolar levels in vitro. In vivo, taxol prevented medial VSMC proliferation and the neointimal VSMC accumulation in the rat carotid artery after balloon dilatation and endothelial denudation injury. This effect occurred at plasma levels approximately two orders of magnitude lower than that used clinically to treat human malignancy (peak levels achieved in this model were approximately 50-60 nM). Taxol may therefore be of therapeutic value in preventing human restenosis with minimal toxicity.

Isoproterenol infusion induces alterations in expression of hypertrophy-associated genes in rat heart

Abstract: Chronic infusion of isoproterenol (Iso) in rats results in cardiac hypertrophy via incompletely understood mechanisms. Our purpose was to determine whether Iso infusion would alter the expression o...

Response of Failing Canine and Human Heart Cells to β2-Adrenergic Stimulation

Abstract: Background Failing human hearts lose β1- but not β2-adrenergic receptors. In canine hearts with tachypacing failure, the ratio of β2- to β1-adrenergic receptors is increased. The present study was designed to determine whether heart failure increases sensitivity to β2-adrenergic stimulation in isolated canine ventricular cardiomyocytes and to verify that myocytes from failing human ventricles contain functional β2-adrenergic receptors. Methods and Results Myocytes from healthy dogs, dogs with tachypacing failure, and human transplant recipients were loaded with fura 2-AM and subjected to electric field stimulation in the presence of zinterol, a highly selective β2-adrenergic agonist. Zinterol significantly increased [Ca2+]i transient amplitudes in all three groups. The failing canine myocytes were significantly more responsive than normal to β2-adrenergic stimulation. We also measured isotonic twitches, indo-1 fluorescence transients, and L-type Ca2+ currents in healthy canine myocytes. Zinterol (10−5 mol...

Role of Calcium/Calmodulin-Dependent Protein Kinase II in the Regulation of Vascular Smooth Muscle Cell Migration

Abstract: Background The migration of vascular smooth muscle cells (VSMCs) is a key event in the pathogenesis of many vascular diseases. We have previously shown that VSMC migration in response to platelet-derived growth factor (PDGF) is suppressed when cultured cells are growth-arrested and induced to differentiate. The present study was undertaken to elucidate the mechanism of this suppression. Methods and Results While both proliferating and growth-arrested VSMCs upregulated expression of the immediate early response genes, c-fos and JE (monocyte chemoattractant protein 1), growth-arrested VSMCs exhibited much smaller changes in intracellular calcium in response to PDGF and failed to activate the calcium/calmodulin-dependent protein kinase II (CaM kinase II). Blocking calcium-calmodulin interactions (50 μmol/L W7) or the activation of CaM kinase II (10 μmol/L KN62) in proliferating cells blocked their migration by more than 90%, whereas inhibition of protein kinase C activation had no significant effect on migra...

Another Modifiable Risk Factor for Cardiovascular Disease? Some Evidence Points to Arterial Stiffness

Abstract: he prevalence of hypertension and atherosclerosis rise T exponentially after age 60, leading to heart failure, stroke, coronary artery disease, and kidney failure and making age the single greatest risk factor for cardiovascular disease (CVD). Why aging, or more specifically why agerelated changes in the heart and arteries, should increase the risk of CVD is a major research question being addressed by the National Institute on Aging (NIA). Many of the cardiovascular changes associated with normal aging have been delineated over the past 20 years. Currently, one area of study focuses on age-related changes in the large arteries and how these changes interact with the pathologies of vascular disease. Increasing evidence points to arterial stiffness-even the amount of stiffness considered normal in western societies-as a critical precursor of disease. Growing insights into the mechanisms underlying stiff ness make it a potential target of interventions, ranging from lifestyle changes to gene therapy, for the prevention and treatment of vascular disease. Hypotheses surrounding arterial stiffness and its relationship to CVD are based partly on the link between stiffening and the age-associated rise in systolic blood pressure. In healthy adults, the walls of the large elastic arteries near the heart increase in both thickness and stiffness with advancing age, and systolic pressure rises. In younger participants in the Baltimore Longitudinal Study of Aging (BLSA), the peak systolic pressure occurs at about the time of peak blood flow. But in older individuals, the stiffening aortic walls speed the pulse waves, which are reflected from the periphery and return to the base of the aorta before the aortic valve closes. The result, a late peak in systolic arterial pressure, is the major determinant of increasing systolic blood pressure with age.’ Various other age-related changes in arteries are associated with stiffening. The amount of elastin relative to collagen in artery walls decreases with age; arterial diameter increases and walls thicken. An increase in vascular tone, determined by increasing calcium concentrations in vascular smooth muscle cells, may also contribute to stiffness. In addition, age-associated stiffening is accompanied by thickening of both the intimal layer of the arterial wall, beneath its inner lining of endothelial cells, and the adjacent medial l a ~ e r . ~ . ~ This thickening of the intima is intriguing because it is caused primarily by an increased number of vascular smooth muscle

The ageing spontaneously hypertensive rat as a model of the transition from stable compensated hypertrophy to heart failure

Abstract: Spontaneously hypertensive rats (SHR) of advanced age exhibit depressed myocardial contractile function and ventricular fibrosis, as stable compensated hypertrophy progresses to heart failure. Transition to heart failure in SHR aged 18-24 months was characterized by impaired left ventricular (LV) function, ventricular dilatation, and reduced ejection fraction without an increase in LV mass. Studies of papillary muscles from SHR with failing hearts (SHR-F), SHR without failure (SHR-NF), and age-matched Wistar Kyoto (WKY) rats allowed examination of changes in the mechanical properties of myocardium during the transition to heart failure. Papillary muscles of SHR-F exhibited increased fibrosis, impaired contraction, and decreased myocyte fractional area. These findings in papillary muscles were correlated with a higher concentration of hydroxyproline and increased histological evidence of fibrosis in the LV free wall. While a depression in active tension accompanied these structural alterations in papillary muscles, it was not evident when active tension was normalized to myocyte fractional area. Together, these data suggest that individual myocyte function may be preserved but that myocyte loss and replacement by extracellular matrix contribute substantially to the decrement in active tension. An absent or negative inotropic response to isoproterenol is observed in SHR-F and SHR-NF papillary muscles and may result in part from age-related alterations in beta-adrenergic receptor dynamics and a shift from alpha- to beta-myosin heavy chain (MHC) protein. During the transition to failure, ventricles of SHR exhibit a marked increase in collagen and fibronectin mRNA levels, suggesting that an increase in the expression of specific extracellular matrix genes may contribute to fibrosis, tissue stiffness, and impaired function. Transforming growth factor-beta 1 (TGF-beta 1) mRNA levels also increase in SHR-F, consistent with the concept that TGF-beta 1 plays a key regulatory role in remodelling of the extracellular matrix gene during the transition to failure. The renin-angiotensin-aldosterone system is also implicated in the transition to failure: SHR treated with the angiotensin converting enzyme inhibitor captopril starting at 12 months of age did not develop heart failure during the 18-24 month observation period. Captopril treatment that was initiated after rats were identified with evidence of failure led to a reappearance of alpha-MHC mRNA but did not improve papillary muscle function. Research opportunities include investigation of apoptosis as a mechanism of cell loss, delineation of the regulatory roles of TGF-beta 1 and the renin-angiotensin-aldosterone system in matrix accumulation, and studies of proteinase cascades that regulate matrix remodelling.

Left Ventricular Diastolic Filling Performance in Older Male Athletes

Abstract: Objective. —To determine whether older men who have undergone intensive endurance training over many years demonstrate less age-associated impairment of early diastolic left ventricular (LV) filling performance than their sedentary peers. Design. —Cross-sectional prospective study. Setting. —Community-dwelling research volunteers. Participants. —Sixteen older competitive male endurance athletes aged 52 through 76 years and 17 young ( Intervention. —All subjects underwent resting Doppler echocardiography and determination of maximal aerobic capacity (Vo 2 max) during graded treadmill exercise. Doppler echocardiographic studies were interpreted without knowledge of the subject's age or exercise habits. Main Outcome Measures. —Doppler-derived measures of LV diastolic filling performance: peak early (E) filling velocity, peak late (A) filling velocity, ratio of peak E to peak A velocities (E/A), and atrial filling fraction. Results. —Older athletes demonstrated higher Vo 2 max (47±6 mL/kg per minute [mean±SD]) than either the young controls (41±7 mL/kg per minute) or older controls (30±7 mL/kg per minute) ( P P P P 2 max was a significant predictor of peak E velocity, peak A velocity, peak E/A ratio, and atrial filling fraction. Conclusion. —Older men with a long history of intensive endurance training demonstrate impaired early diastolic LV filling similar to that of their sedentary peers. Thus, impairment of early diastolic filling appears to be intrinsic to normative aging and not secondary to the reduction in aerobic capacity that accompanies the aging process. ( JAMA . 1995;273:1371-1375)

Intracellular signaling pathways required for rat vascular smooth muscle cell migration. Interactions between basic fibroblast growth factor and platelet-derived growth factor.

Abstract: Intracellular signaling pathways activated by both PDGF and basic fibroblast growth factor (bFGF) have been implicated in the migration of vascular smooth muscle cells (VSMC), a key step in the pathogenesis of many vascular diseases. We demonstrate here that, while bFGF is a weak chemoattractant for VSMCs, it is required for the PDGF-directed migration of VSMCs and the activation of calcium/calmodulin-dependent protein kinase II (CamKinase II), an intracellular event that we have previously shown to be important in the regulation of VSMC migration. Neutralizing antibodies to bFGF caused a dramatic reduction in the size of the intracellular calcium transient normally seen after PDGF stimulation and inhibited both PDGF-directed VSMC migration and CamKinase II activation. Partially restoring the calcium transient with ionomycin restored migration and CamKinase II activation as did the forced expression of a mutant CamKinase II that had been "locked" in the active state by site-directed mutagenesis. These results suggest that bFGF links PDGF receptor stimulation to changes in intracellular calcium and CamKinase II activation, reinforcing the central role played by CamKinase II in regulating VSMC migration.

Effects of sarcoplasmic reticulum Ca2+ load on the gain function of Ca2+ release by Ca2+ current in cardiac cells

Abstract: We studied the effects of variable sarcoplasmic reticulum (SR) Ca2+ loading on changes in the gain index of Ca2+ release from the SR, measured as the ratio of the amount of Ca2+ released to the magnitude of the Ca2+ current (ICa) integrated for the initial 20 ms of the depolarization, in whole cell voltage-clamped rat ventricular myocytes dialyzed with the Ca2+ indicator indo 1 salt at 23 degrees C. Changes in ICa were measured directly, and changes in the SR Ca2+ release were indexed by changes in the amplitudes and rates of rise of cytosolic Ca2+ (Ca2+i) transients. The SR Ca2+ load was graded by the duration of conditioning voltage-clamp steps and verified by caffeine-dependent Ca2+i transients. A train of abbreviated (from 100 to 20 ms) voltage-clamp depolarizations, which triggers SR Ca2+ release but fails to replenish the SR with Ca2+, diminished the SR Ca2+ load by 56 +/- 5%, did not alter peak ICa but reduced the amplitudes of the ICa-dependent Ca2+i transients by 52 +/- 3%, and decreased the gain index by 60 +/- 3% (SE; n = 5 or 6). Changes in the amplitudes of Ca2+i transients elicited by ICa and changes in the gain index were linearly correlated (r2 = 0.83 and 0.79, respectively; P < 0.001 for each) with changes in amplitudes of Ca2+i transients elicited by caffeine pulses applied in lieu of the respective voltage-clamp pulses.(ABSTRACT TRUNCATED AT 250 WORDS)

Age-associated increase in rat ventricular ANP gene expression correlates with cardiac hypertrophy.

Abstract: Atrial natriuretic peptide (ANP), a cardiac-specific hormone, is stored in the atria and released in response to atrial stretch. During cardiac hypertrophy, ANP gene expression is markedly upregulated in the left ventricle (LV). Because the hearts of normotensive senescent rats exhibit left atrial (LA) and left ventricular (LV) hypertrophy and dilatation, we examined ANP mRNA levels by Northern blot analysis and ANP peptide concentrations by radioimmunoassay in atria, LVs, and plasma of rats at 2, 6, 18, and 22-24 mo of age. Compared with LVs of 6-mo-old rats, the LV-to-body weight ratio was elevated 30% by 18 mo of age, whereas levels of ANP mRNA were elevated twofold (not significant) and sevenfold (P < 0.05) in the LV of 18- and 22- to 24-mo-old rats, respectively. The concentration of immunoreactive ANP (ir-ANP) exhibited a four- to fivefold increase in LVs of 18- and 22- to 24-mo-old rats compared with values for 6-mo-old rats (43 +/- 4 pmol/g wet wt; means +/- SE). Among 18-and 22- to 24-mo-old rats a significant correlation was observed between ANP peptide concentration and LV hypertrophy (r 2 = 0.64). Levels of ANP mRNA and ir-ANP in the atria exhibited only modest changes with aging.(ABSTRACT TRUNCATED AT 250 WORDS)

Reduced threshold for myocardial cell calcium intolerance in the rat heart with aging.

Abstract: To determine whether advancing age is accompanied by a reduced Ca2+ tolerance, we measured Ca(2+)-dependent diastolic pressure, prolonged relaxation and systolic functional deterioration, spontaneous sarcoplasmic reticulum (SR)-generated Ca2+ oscillations [detected as scattered laser light intensity fluctuations (SLIF)], aftercontractions, and ventricular fibrillation in isolated, isovolumic, atrioventricular-blocked intact hearts from 24- to 26-mo (old) and 6- to 8-mo (young) male Wistar rats. In enzymatically isolated single cardiac myocytes, the likelihood of the occurrence of spontaneous contractile waves driven by spontaneous SR Ca2+ release was also determined. In response to stepwise increase in perfusate Ca2+ concentration (Cao), a reduction in the maximum developed pressure accompanied by an elevation in end-diastolic pressure and a prolonged contraction duration was observed at lower Cao in old vs. young hearts (P < 0.01 for each parameter). Furthermore, Ca(2+)-dependent ventricular fibrillation occurred during pacing in six old but in no young hearts (P < 0.01), aftercontractions were observed in seven old vs. one young heart (P < 0.01), and SLIF increased to a greater extent in old vs. young hearts. In single cardiac myocytes, spontaneous contractile waves occurred more frequently with increasing age (P < 0.01). These results indicate that aging is associated with an increased likelihood for the occurrence of SR-generated Ca2+ oscillations and functional abnormalities that result from these oscillations.

Endoplasmic Reticulum Ca2+ Depletion Unmasks a Caffeine-Induced Ca2+ Influx in Human Aortic Endothelial Cells

Abstract: Intracellular Ca2+ pools contribute to changes in cytosolic [Ca2+] ([Ca2+]i), which play an important role in endothelial cell signaling. Recently, endothelial ryanodine-sensitive Ca2+ stores were shown to regulate agonist-sensitive intracellular Ca2+ pools. Since caffeine binds the ryanodine Ca2+ release channel on the endoplasmic reticulum in a variety of cell types, we examined the effect of caffeine on [Ca2+]i in human aortic endothelial cell monolayers loaded with the fluorescent probe indo 1. Under baseline conditions, 10 mmol/L caffeine induced a small increase in [Ca2+]i from 86±10 to 115±17 nmol/L (mean±SEM); this effect was similar to that of 5 μmol/L ryanodine and was unaffected by buffer Ca2+ removal. After depletion of an intracellular Ca2+ store by the irreversible endoplasmic reticulum Ca2+-ATPase inhibitor thapsigargin (1 μmol/L), ryanodine did not affect [Ca2+]i. In contrast, caffeine induced a large rapid increase in [Ca2+]i (176±19 to 338±35 nmol/L, P<.001) after thapsigargin e...

cGMP prevents delayed relaxation at reoxygenation after brief hypoxia in isolated cardiac myocytes

Abstract: Previous studies in isolated cardiac myocytes suggest that impaired relaxation during reoxygenation after brief hypoxia results from abnormal Ca(2+)-myofilament interaction. Recent studies indicate that guanosine 3',5'-cyclic monophosphate (cGMP)-elevating interventions selectively enhance myocardial relaxation. We investigated the effect of 8-bromoguanosine 3',5'-cyclic monophosphate (8-BrcGMP) on posthypoxic relaxation in single rat myocytes, with simultaneous measurement of contraction and intracellular Ca2+ (indo 1 fluorescence). In control myocytes (n = 11), reoxygenation after 10 min of hypoxia markedly prolonged time to peak shortening (+36.5 +/- 4.2%) and half-relaxation time (+75.7 +/- 11.3% cf. normoxic values; both P < 0.001) and reduced diastolic length but did not change cytosolic Ca2+. Under normoxic conditions, 50 microM 8-BrcGMP slightly reduced time to peak shortening and half-relaxation time and increased diastolic length but did not alter cytosolic Ca2+. In the presence of 8-BrcGMP, there was no posthypoxic delay in twitch relaxation nor was there a decrease in diastolic length (half-relaxation time -5.8 +/- 3.3% cf. normoxic values; P < 0.05 cf. control group; n = 11). Cytosolic Ca2+ remained unaltered. Thus, 8-BrcGMP fully prevents impaired posthypoxic relaxation in isolated cardiac myocytes, probably by altering Ca(2+)-myofilament interaction.

cGMP prevents delayed relaxation at reoxygenation after bried hypoxia in isolated cardiac myocytes

Abstract: Previous studies in isolated cardiac myocytes suggest that impaired relaxation during reoxygenation after brief hypoxia results from abnormal Ca 2+ -myofilament interaction. Recent studies indicate that guanosine 3',5'-cyclic monophosphate (cGMP)-elevating interventions selectively enhance myocardial relaxation. We investigated the effect of 8-bromoguanosine 3',5'-cyclic monophosphate (8-BrcGMP) on posthypoxic relaxation in single rat myocytes, with simultaneous measurement of contraction and intracellular Ca 2+ (indo 1 fluorescence). In control myocytes (n = 11), reoxygenation after 10 min of hypoxia markedly prolonged time to peak shortening (+ 36.5 ± 4.2%) and half-relaxation time (+75.7 ± 11.3% cf. normoxic values ; both P < 0.001) and reduced diastolic length but did not change cytosolic Ca 2+ . Under normoxic conditions, 50 μM 8-BrcGMP slightly reduced time to peak shortening and half-relaxation time and increased diastolic length but did not alter cytosolic Ca 2+ . In the presence of 8-BrcGMP, there was no posthypoxic delay in twitch relaxation nor was there a decrease in diastolic length (half-relaxation time -5.8 ± 3.3% cf. normoxic values ; P < 0.05 cf. control group ; n = 11). Cytosolic Ca 2+ remained unaltered. Thus, 8-BrcGMP fully prevents impaired posthypoxic relaxation in isolated cardiac myocytes, probably by altering Ca 2+ -myofilament interaction.

The Sustained Release of Galardin and Taxol from Gelatin Chondroitin Sulfate Coacervate Films

Abstract: Coronary artery atherosclerosis is the leading cause of death in the United States. Restenosis following percutaneous transluminal balloon angioplasty (PTCA) remains the limiting factor in the use of this treatment for coronary artery disease. Restenosis occurs in 30% of patients within 6 months. The restenotic lesion is a fibroproliferative response with resulting smooth muscle cell migration, proliferation and extracellular matrix production. Despite a decade of research there is no effective strategy for preventing restenosis in man.