摘要:

Phospholamban is the regulator of the Ca2+-ATPase in cardiac sarcoplasmic reticulum (SR), and it has been suggested to be an important determinant in the inotropic responses of the heart to β-adrenergic stimulation. To determine the role of phospholamban in vivo, the gene coding for this protein was targeted in murine embryonic stem cells, and mice deficient in phospholamban were generated. The phospholamban-deficient mice showed no gross developmental abnormalities but exhibited enhanced myocardial performance without changes in heart rate. The time to peak pressure and the time to half-relaxation were significantly shorter in phospholamban-deficient mice compared with their wild-type homozygous littermates as assessed in work-performing mouse heart preparations under identical venous returns, afterloads, and heart rates. The first derivatives of intraventricular pressure (±dP/dt) were also significantly elevated, and this was associated with an increase in the affinity of the SR Ca2+-ATPase for Ca2+in the phospholamban-deficient hearts. Baseline levels of these parameters in the phospholamban-deficient hearts were equal to those observed in hearts of wild-type littermates maximally stimulated with the β-agonist isoproterenol. These findings indicate that phospholamban acts as a critical repressor of basal myocardial contractility and may be the key phosphoprotein in mediating the heart's contractile responses to β-adrenergic agonists.

ISSN:0009-7330    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

Previous studies have indicated the focal presence of growth factors and focal low levels of cell proliferation in human atherosclerotic plaques. Using human carotid plaques and an antibody to platelet-derived growth factor (PDGF)-A chain, we have begun to assess growth factor significance by spatially correlating growth factor gene expression with actual cell proliferation. Since PDGF is a mitogen for smooth muscle and related cells and since inflammatory cells (eg, macrophages) can also proliferate in these lesions, it was important to exclude inflammatory cell proliferation from this consideration. Therefore, we have used a triple immunolabeling approach, combining the above anti–PDGF-A chain antibody with an inflammatory cell cocktail (CD68+CD45 for monocyte/macrophages and lymphocytes) and adding an anti-proliferating cell nuclear antigen (PCNA) antibody to mark proliferating cells. In the carotid atherosclerotic plaques, PDGF immunostaining was distributed focally, preferentially in the fibrous cap and vascularized regions, and was present in two distinct patterns: cytoplasmic and diffuse extracellular staining. When we considered colocalization within the same cells, cytoplasmic PDGF-A staining did not appear to colocalize with inflammatory markers. PCNA nuclear staining combined with PDGF cytoplasmic staining of the same cell was detected extremely rarely. Considering colocalization within the same microscopic fields, PDGF-A staining was detected more frequently than noninflammatory PCNA positivity. Quantitative logistic regression analysis demonstrated that localization in vascularized regions and (independently) the presence of PDGF-A are good predictors of noninflammatory cell proliferation, within the same microscopic fields. Therefore, PDGF-A and other factors especially associated with vascularized regions may be involved in the regulation of mesenchymal cell proliferation in human atherosclerotic plaques.

ISSN:0009-7330    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

The left ventricular hypertrophy that develops with the volume overload of mitral regurgitation is relatively less than that which develops with the pressure overload of aortic stenosis even when both lesions are severe. The hypertrophy that develops must be the sum of changes in the rate of myocardial protein synthesis and degradation. In the present canine study, we explored early changes in the synthesis rate of myosin heavy chain in response to severe acute pressure overload versus that of the severe acute volume overload of mitral regurgitation. We tested the hypothesis that in acute overload, the rate of protein synthesis would increase less in the volume-overload model than in the pressure-overload model, a potential partial mechanism for the discrepancy in the eventual total amount of hypertrophy that develops in these two lesions. Acute pressure overload was produced by inflating a balloon in the descending aorta, and acute volume overload was produced by using our closed-chest mitral chordal rupture technique. In both models, the hemodynamic lesion that was created was severe. In eight dogs with pressure overload, the average gradient across the balloon was 119.8±6.1 mm Hg. In six dogs with volume overload, the average regurgitant fraction was 0.67±0.06. Six other dogs served as controls. The average rate of myosin heavy chain synthesis in control dogs was 2.7±0.2% per day, virtually identical to the rate we found in the severe volume-overload model. In contrast, the rate was increased in the pressure-overload model by 30% to 3.5±0.3% per day (P<.05). We conclude that the rate of myocardial protein synthesis increases measurably after 6 hours of severe pressure overload but does not after 6 hours of severe volume overload. If these early qualitative differences persisted, they would help explain the relative lack of hypertrophy in the volume overload of mitral regurgitation.

ISSN:0009-7330    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

We examined whether apoptosis occurs in cardiomyocytes by hypoxia in vitro. Neonatal rat cardiomyocytes and nonmyocytes were cultured in 95% N2–5% CO2atmosphere to produce hypoxic conditions. DNA fragmentation into integer multiples of the internucleosomal DNA length was observed in cardiomyocytes as early as 12 hours, whereas nonmyocytes did not show fragmentation of DNA up to 72 hours. DNA fragmentation of cardiomyocytes induced by hypoxia was also confirmed by nick-end labeling in situ. Messenger RNA for Fas antigen, a mediator of apoptotic cell death, was expressed in both cardiomyocytes and nonmyocytes as revealed by Northern blotting and in situ hybridization. In hypoxic conditions, Fas messenger RNA levels in cardiomyocytes were upregulated by twofold over controls, whereas those of nonmyocytes were downregulated. These results indicate that cardiomyocyte death by hypoxia can occur via apoptosis and that Fas antigen may be associated with the mechanism of this apoptotic process.

ISSN:0009-7330    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

Expression of sarcoplasmic reticulum (SR) Ca2+-ATPase was shown to be reduced in failing human myocardium. The functional relevance of this finding, however, is not known. We investigated the relation between myocardial function and protein levels of SR Ca2+-ATPase in nonfailing human myocardium (8 muscle strips from 4 hearts) and in myocardium from end-stage failing hearts with dilated (10 muscle strips from 9 hearts) or ischemic (7 muscle strips from 5 hearts) cardiomyopathy. Myocardial function was evaluated by the force-frequency relation in isometrically contracting muscle strip preparations (37°C, 30 to 180 min−1). In nonfailing myocardium, twitch tension rose with increasing rates of stimulation and was 76% higher at 120 min−1compared with 30 min−1(P<.02). In failing myocardium, there was no significant increase in average tension at stimulation rates above 30 min−1. At 120 min−1, twitch tension was decreased by 59% (P<.05) in dilated cardiomyopathy and 76% (P<.05) in ischemic cardiomyopathy compared with nonfailing myocardium. Protein levels of SR Ca2+-ATPase, normalized per total protein or per myosin, were reduced by 36% (P<.02) or 32% (P<.05), respectively, in failing compared with nonfailing myocardium. SR Ca2+-ATPase protein levels were closely related to SR Ca2+uptake, measured in homogenates from the same hearts (r=.70, n=16, andP<.005). For all types of myocardium, there was a significant correlation between SR Ca2+-ATPase protein levels and the frequency at which twitch tension was maximum (r=.74, n=18, andP<.001 in the case of normalization per total protein). Similarly, SR Ca2+-ATPase protein levels were correlated with the potentiation of twitch tension after an increase in stimulation frequency from 30 to 120 min−1(r=.80, n=18, andP<.001). These data show that the protein levels of SR Ca2+-ATPase are closely related to the force-frequency behavior of human myocardium. This may suggest that protein levels of SR Ca2+-ATPase determine the systolic contractile reserve with respect to frequency potentiation of contractile force in the human myocardium.

ISSN:0009-7330    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

The regulation of cytosolic Ca2+concentration during excitation-contraction coupling is altered in the failing human heart. Previous studies have focused on disturbances in Ca2+release and reuptake from the sarcoplasmic reticulum (SR), whereas functional studies of the cardiac Na+-Ca2+exchanger, another important determinant of myocyte homeostasis, are lacking for the failing human heart. Using a cardiac Na+-Ca2+exchanger cDNA recently cloned from a guinea pig cDNA library, we investigated the gene expression of the cardiac Na+-Ca2+exchanger in relation to the SR Ca2+-ATPase. Expression of both genes was quantified in left ventricular myocardium from 24 failing human cardiac explants and 7 control heart samples in relation to β-myosin heavy chain mRNA by slot blot analysis. Compared with patients with nonfailing hearts, patients with dilated cardiomyopathy (DCM, n=13) showed a 55% increase in Na+-Ca2+exchanger mRNA levels (P<.05 versus control value) and a 41% increase in patients with coronary artery disease (CAD, n=11). In the same hearts, SR Ca2+-ATPase mRNA levels were decreased by 50% in DCM and by 45% in CAD (P<.05 for both versus control value). There was a positive correlation between Na+-Ca2+exchanger and SR Ca2+-ATPase mRNA levels both in normal and failing human hearts, albeit with different slopes and intercepts of the regression line. The Na+-Ca2+exchanger protein levels as assessed by Western blot analysis and normalized to β-myosin heavy chain protein were increased in DCM and CAD (P<.05 andP<.01 versus control value, respectively), whereas SR Ca2+-ATPase protein levels were reduced (P<.05 for both groups versus control values). Thus, the Na+-Ca2+exchanger gene expression is enhanced in failing human hearts and may, in part, compensate for the depressed SR function with regard to diastolic Ca2+removal.

ISSN:0009-7330    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

The renin-angiotensin and cardiac natriuretic systems play an important role in the pathophysiology of congestive heart failure (CHF). The status of the membrane-bound pulmonary and renal activities of three ectoenzymes involved in the regulation of these systems — angiotensin-converting enzyme (ACE), neutral endopeptidase (NEP), and aminopeptidase A (APA) —was investigated in Wistar rats 3 months after induction of myocardial infarction (MI) and in sham-operated (control) rats. Plasma renin activity and ACE activity, plasma angiotensin II (Ang II) levels, and atrial natriuretic factor levels were simultaneously determined. The lung ACE activity was decreased in MI rats compared with control rats (P<.0001), and this decrease depended on the severity of the heart failure. In contrast, plasma ACE activity was increased in MI rats (P<.01), and this increase was also proportional to the severity of MI. Northern blot analysis showed that the lung ACE mRNA level in severe MI rats was half that of the control rats. Renal ACE activity of the MI rats was not affected, and neither renal or pulmonary NEP nor pulmonary APA activities were altered. Thus, lung ACE gene expression appears to be both organ- and enzyme-specifically regulated during CHF. Whereas plasma renin was increased in heart failure rats, plasma Ang II levels were not different from those of control rats. Thus, decreased lung ACE activity could possibly contribute to keeping plasma Ang II levels in the normal range. The decrease in lung ACE activity and mRNA levels, combined with increased plasma ACE activity, represents a novel aspect of endothelial dysfunction in CHF. This dissociation between the membrane-bound endothelial enzyme and its circulating counterpart emphasizes the importance of simultaneously assessing the circulating and tissue components of the renin-angiotensin system in heart failure.

ISSN:0009-7330    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

We developed a mouse myocardial preparation to study cellular dysfunction in acute coxsackievirus myocarditis. Thin right ventricular papillary muscles from normal mice (n=8) were compared with muscles from mice 7 days after coxsackievirus infection (n=7). Sarcomere shortening was studied with laser diffraction (HeNe, λ=623.8 nm). A servomotor was used to shorten a muscle until slack early in isometric contraction. Unloaded sarcomere shortening velocity (Vo) was measured at the start of zero force at slack length. Vowas independent of the extent of slack release and was the same as that estimated with an isotonic force-sarcomere shortening velocity relation. Resting muscle stiffness was calculated from shortening perturbations in resting muscles. The histology of some papillary muscles (normal, n=4; infected, n=3) was studied. There was no ventricular hypertrophy. Resting sarcomere length (SL) in infected preparations (2.11±0.07 μm) (mean±1 SD) was the same as in normal preparations (2.11±0.08 μm). In isometric twitches in normal and infected muscles, total peak force (4.31±1.07 and 3.77±1.86 g/mm2, respectively) resting force (0.81±0.37 and 0.81±0.35 g/mm2, respectively), and time to peak force (129.5±20.3 and 125.2±13.0 milliseconds, respectively) were not significantly different. Vowas 4.14±0.84 μm/s in normal muscles at an SL of 2.08±0.09 μm and 1.70±0.33 μm/s in infected muscles at an SL of 2.06±0.08 μm. Resting stiffness was the same for normal and infected muscles. There was inflammation but no fibrosis or necrosis. Thus, Vowas depressed early in acute viral myocarditis without hypertrophy, myocyte necrosis, fibrosis, or altered resting stiffness. Pyrophosphate gel electrophoresis showed a shift from predominantly fast to slow myosin isoforms. Apparently, there is remodeling of the contractile apparatus early in acute coxsackievirus myocarditis that is caused either by the direct effects of the virus or the immune response.

ISSN:0009-7330    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

In various mammalian species, shapes of action potentials vary within the cardiac wall because of differences in transient outward current (Ito). A prominent Itoexists in human ventricular myocytes, but cells have not been separated according to their original localization. Human ventricular myocytes were isolated from separated subepicardial and subendocardial tissue, and regional variations in Itowere studied. Itowas larger in subepicardial than subendocardial cells. Current density at +60 mV was 7.9±0.7 pA/pF (n=28) in subepicardial cells and 2.3±0.3 pA/pF (n=16) in subendocardial cells. When cells from explanted failing and nonfailing donor hearts were compared, Itowas not different in subepicardial cells; however, it was larger in subendocardial cells from nonfailing hearts. The potential of half-maximal activation (V0.5) was more positive in subendocardial cells (+25.6±3.5 mV, n=15) than in subepicardial cells (+9.2±1.8 mV, n=28). There was no difference in V0.5between cells from failing and nonfailing hearts. Itoinactivation was similar in all cell types and independent of membrane depolarization (time constant [τ]=≈60 milliseconds at 22°C). The potential of half-maximal steady-state inactivation was similar in all cell types. Recovery from inactivation of Itowas fast in subepicardial cells at −100 mV (τ=24±4 milliseconds, n=6), exceeding control values transiently (overshoot), and slow at −40 mV without overshoot (τ=638±91 milliseconds, n=6). In subendocardial cells, Ito, recovered at −100 mV with a fast phase (τ=25 milliseconds) and a slow phase (τ=328 milliseconds), and recovery was not complete after 6 seconds at −100 mV. In conclusion, regional differences in Itobetween subepicardial and subendocardial cells may have clinical implications with respect to rhythmic disturbance during heart failure.

ISSN:0009-7330    

出版商:OVID    

年代:1994

数据来源: OVID

摘要:

Gap junctions are thought to mediate the direct intercellular coupling of adjacent cells by the gating of an aqueous pore permeable to ions and molecules of up to 1 kD or 8 to 14 Å in diameter. We performed ion-substitution and dye-transfer experiments to determine the relative Cl−/K+conductance and dye permeability of anionic fluorescein derivatives in chick connexin45 (Cx45) channels. We demonstrate that Cx45 forms a 26±6-picosiemen (pS) channel with a maximum detectable Cl−permeability of 0.2 relative to K+or Cs+. Although homogeneous channel conductances were observed in multichannel recordings, the open probability estimates were indicative of nonhomogeneous gating behavior and occasional cooperativity. A second conductance state of 19±4 pS begins to predominate at higher voltages. Cx45 gap junctions are permeable to 2′,7′-dichlorofluorescein but are not permeable to the more polar 6-carboxyfluorescein dye. These observations suggest that the Cx45 pore diameter is ≈ 10 Å and is associated with a fixed negative charge within the junctional channel.

ISSN:0009-7330    

出版商:OVID    

年代:1994

数据来源: OVID