Pulsatile hemodynamics in congestive heart failure.

Pulse pressure, an indirect measure of vascular stiffness and pulsatile load, predicts clinical events in congestive heart failure (CHF), suggesting that abnormal pulsatile load may contribute to CHF. This study was designed to assess more direct measures of central pulsatile load in CHF. Noninvasive hemodynamic evaluations were performed in 28 subjects with CHF and 40 controls using calibrated tonometry of the brachial, radial, femoral, and carotid arteries along with echocardiographic assessment of left ventricular outflow tract (LVOT) diameter and Doppler flow. Characteristic impedance (Z(c)) was calculated as the ratio of DeltaP (carotid) and DeltaQ (LVOT flow) in early systole. Carotid-radial (CR-PWV) and carotid-femoral (CF-PWV) pulse wave velocities were calculated from tonometry. Augmentation index was assessed from the carotid waveform. Total arterial compliance (TAC) was calculated using the area method. Brachial pulse pressure was elevated (62+/-16 versus 53+/-15 mm Hg, P=0.015) in CHF because of lower diastolic pressure (66+/-10 versus 73+/-9 mm Hg, P=0.003). CHF had higher Z(c) (225+/-76 versus 184+/-66 dyne. sec. cm(-5), P=0.020). CF-PWV did not differ (9.7+/-2.7 versus 9.2+/-2.0, P=0.337), whereas CR-PWV was lower in CHF (8.6+/-1.4 versus 9.4+/-1.5, P=0.038). There was no difference in TAC (1.4+/-0.5 versus 1.4+/-0.6 mL/mmHg, P=0.685), and augmentation index was lower in CHF (8+/-17 versus 21+/-13%, P=0.001). CHF subjects have elevated central pulsatile load (Z(c)), which is not apparent in global measures such as augmentation index or TAC, possibly because of contrasting changes in central and peripheral conduit vessels. This increased pulsatile load represents an important therapeutic target in CHF.

Genetic mechanisms of sudden cardiac death.

When there is a genetic mutation in a major cardiac protein, SCD is often an unfortunate part of the clinical phenotype. In the majority of common cardiac diseases, even though the nature or degree of genetic influence is not understood their importance cannot be denied. Although advancing by one gene or gene interaction at a time is arduous, it is the only apparent way to begin to unravel as complex a clinic syndrome as SCD.

Recent advances in dobutamine stress echocardiography.

Dobutamine stress echocardiography (DSE) is a reliable cardiac risk stratifier that has widespread applicability because of its clinical accuracy and cost effectiveness. Dobutamine has positive inotropic and chronotropic effects and is commonly used in patients who cannot exercise or achieve an adequate heart rate response with exercise. Recently available long-term results from several independent clinical trials, combined with enhancements in image quality, have improved the ability to detect significant coronary artery disease and determine myocardial viability. Dobutamine stress echocardiography has an excellent safety profile with clinical results superior to regular exercise electrocardiography and comparable with exercise echocardiography and radionucleotide perfusion stress imaging. Low-dose dobutamine response can accurately predict dysfunctional yet viable myocardial regions that may improve with revascularization. Clinical studies are now available refining the common use of DSE preoperatively in female patients with valvular disease, as well as in the emergency department. Dobutamine stress echocardiography does have some limitations in discriminating particular regions of ischemia when multiple ventricular segments are involved and when the imaging is suboptimal. It can be applied using minimal additional resources in an otherwise functioning echocardiography laboratory and, with appropriate training, can result in clinical results comparable with those of large-scale multicenter trials. Ongoing improvements in technology and the development of new reagents such as myocardial contrast agents hold promise for further advancement in the near future.

Upcoming therapies for heart failure.

Treatment of maladaptive neurohormonal activation in congestive heart failure (CHF) has been successful because basic cardiovascular science findings have been confirmed or dismissed through the use of well-controlled, large-scale clinical trials. It should be no surprise that this exciting approach is evolving toward novel agents and devices directed toward other pathways involved in CHF neurohormonal/cytokine activation. Several of these are in advanced clinical development and are likely to play prominent roles in CHF therapy in the near future.

F1-ATP synthase beta-subunit and cytochrome c transcriptional regulation in right ventricular hemodynamic overload and hypertrophically stimulated cardiocytes.

Cardiac hypertrophic growth secondary to hemodynamic pressure overload causes changes in energy requirements that may involve the transcriptional upregulation of oxidative phosphorylation genes. Therefore, two representative nuclear-encoded genes, the mitochondrial F1-ATP synthase beta-subunit (beta-subunit) and cytochrome c (cyt c), were examined in a feline chronic pulmonary artery banded right ventricular pressure-overload model. In the hypertrophying right ventricle, beta-subunit and cyt c mRNA levels increased after two and seven days, during the peak growth response. To examine cardiac transcriptional regulation, neonatal rat cardiac myocytes (cardiocytes) were transiently transfected with beta-subunit promoter constructs ranging from -1519 nucleotides (nt) upstream of transcription initiation as well as cyt c promoter constructs ranging from -726 nt. A full-length p1519beta-subunit/Luc construct was alpha-adrenergically inducible by 275% (+/-30%) with this activation being mapped to an enhancer region between -1519 to -1480 nt. Smaller constructs containing more proximal promoter elements were not inducible. Additionally, the full-length and enhancer deleted beta-subunit constructs were also inducible in electrically stimulated cardiocytes, suggesting a different mechanism of activation. Cyt c constructs containing known constitutive elements from -191 to -167 nt and -139 to -84 nt were responsible for the majority of the reporter activity of the full-length promoter but were not inducible in the presence of phenylephrine. Hence, we show that promoter regions containing elements common in other metabolism-related gene families are active in neonatal rat cardiocytes. Once more, we have identified a beta-subunit genomic region responsive to alpha-adrenergic and electrical stimulation.

Upregulation of the cardiac homeobox gene Nkx2-5 (CSX) in feline right ventricular pressure overload.

The recent characterization of the cardiac-specific homeobox gene Nkx2-5 (or CSX) and its detection in normal adult heart tissue raises the possibility of a role in adult hypertrophy. Using pressure overload as a primary stimulus, we used a feline pulmonary artery banding model to produce right ventricular hypertrophy (RVH). Total RNA was hybridized to a full-length murine Nkx2-5 cDNA probe that contained the NK family homeodomain. Nkx2-5 mRNA levels increased 5.1-fold (P < 0.05) and 3.9-fold vs. the corresponding left ventricles at 2 and 7 days of RVH, respectively, during the period of maximal myocardial growth. By 2 wk, when the RVH response had been completed, Nkx2-5 mRNA levels were returning toward baseline. Hybridization with an Nkx2-5 probe not containing the NK homologous homeodomain demonstrated that upregulation was specific for the Nkx2-5 gene. Atrial natriuretic factor and alpha-cardiac actin, both activated in part by Nkx2-5 DNA binding elements, also increased with RVH. These data suggest that a cardiac homeobox gene may play a role in the induction of adult cardiac hypertrophy.

Chamber specification of atrial myosin light chain-2 expression precedes septation during murine cardiogenesis.

To study the molecular mechanisms that control patterning of the heart tube during early cardiogenesis, we have used the ventricular myosin regulatory light chain (MLC-2v), which is expressed in the ventricular segment of the primitive heart tube, as a genetic marker for ventricular specification in rodents. To assess whether the atrial isoform, MLC-2a, could also serve as a chamber-specific marker, we cloned an atrial MLC-2 cDNA (554 base pairs) which displayed homology to the human MLC-2a cDNA at both the nucleotide (87%) and amino acid (95%) levels. Northern blot, reverse transcriptase-linked polymerase chain reaction, RNase protection, and Western blot analysis revealed atrial restricted expression in the adult mouse heart, very low levels in aorta, and no detectable expression in ventricle, skeletal muscle, uterus, or liver. In situ hybridization studies during mouse embryogenesis revealed cardiac specific expression throughout days 8-16 postcoitum, with atrial restricted expression from day 12 and qualitatively greater atrial expression than ventricular from day 9. Thus, preferential pattern of expression in the atria occurs prior to septation. The MLC-2a gene was differentially regulated when compared with MLC-2v expression during embryonic stem cell cardiogenesis in vitro with MLC-2a transcript levels detectable from day 6 in suspension cultures as compared with day 9 for MLC-2v. The region-specific expression of the MLC-2a and MLC-2v genes in their respective chambers during early cardiogenesis provides genetic markers for chamber specification (atrial and ventricular) in both the in vitro and in vivo context.

Expression of the helix-loop-helix factor Id during mouse embryonic development.

Expression and regulation of the helix-loop-helix (HLH) protein Id in cardiac cells and its negative effect on cardiac gene expression indicate that Id may play a role in cardiac development. To investigate this issue further, we have performed in situ analysis of Id mRNA in developing mouse embryos from Embryonic Days 7 to 15.5 and in neonatal heart. Consistent with its postulated role as an inhibitor of differentiation, Id mRNA was expressed in specific populations of proliferating mesenchymal and epithelial tissues, with its expression declining as differentiation progressed. In addition, patterns of Id mRNA hybridization overlapped considerably with those of two other developmentally important genes, twist and hox 7/7.1, of the HLH and homeobox family, respectively. Id mRNA hybridization was not observed in early epithelial somites or in the myotomal compartment of the somite, suggesting that Id is unlikely to play an inhibitory role in these early myogenic precursors. Id mRNA was expressed in specific regions of the developing nervous system. In the heart, Id mRNA was expressed at high levels in developing cardiac cushions and ridges of the outflow tract and continued to be expressed at high levels in valvular tissue of the neonatal heart.

Positional specification of ventricular myosin light chain 2 expression in the primitive murine heart tube.

To study the process of ventricular specification during cardiogenesis, we examined the in situ expression of cardiac ventricular myosin light chain 2 (MLC-2v) mRNA during murine embryogenesis. As assessed by hybridization with a specific MLC-2v riboprobe, mRNA expression can be found in the ventricular region at day 8.0 postcoitum (pc). MLC-2v expression is high in the ventricular portion of the heart tube, with no detectable expression in the atrial or sinus venosus regions. The proximal outflow tract of the heart tube also expresses MLC-2v mRNA at minimally detectable levels at this time but then displays a temporally and spatially distinct pattern with expression well established in the proximal out-flow tract region adjacent to the ventricular segment by days 9-10 pc, eventually reaching levels comparable to the trabeculated ventricular myocardium. By day 11 pc, prior to the completion of septation, expression then becomes restricted to the ventricular region at and below the level of the atrioventricular cushion. Transgenic mice harboring a 250-base-pair MLC-2v promoter fragment fused to a luciferase reporter gene demonstrate reporter gene activity from at least day 9 pc. Ventricular region-restricted expression of the luciferase reporter in the embryonic heart, as assessed by immunofluorescence and direct assay of reporter activity in microdissected atrial and ventricular muscle specimens, was confirmed from at least day 15 pc on. Taken together, this provides evidence for early positional specification of MLC-2v gene expression in the primitive heart tube and indicates regional specification of part of the ventricular muscle gene program can precede ventricular septation during mammalian cardiogenesis. Since the 250-base-pair promoter fragment is active developmentally in transgenic mice, this establishes it as a molecular target for the process of ventricular specification in the developing heart tube.

Myosin light chain-2 luciferase transgenic mice reveal distinct regulatory programs for cardiac and skeletal muscle-specific expression of a single contractile protein gene.

To examine the relationship between the cardiac and skeletal muscle gene programs, the current study employs the regulatory (phosphorylatable) myosin light chain (MLC-2) as a model system. Northern blotting, primer extension, and RNase protection studies documented the high level expression of the cardiac MLC-2 mRNA in both mouse cardiac and slow skeletal muscle (soleus). Transgenic mouse lines harboring a 2100- or a 250-base pair rat cardiac MLC-2 promoter/luciferase fusion gene were generated, demonstrating high levels of luciferase activity in cardiac muscle, and only background luminescence in slow skeletal muscle and non-muscle tissues. As assessed by in situ hybridization, immunofluorescence, and luminescence assays of luciferase reporter activity in various regions of the heart, both the endogenous MLC-2 gene and the MLC-2 luciferase fusion gene were expressed exclusively in the ventricular compartment, with expression in the atrium at background levels. Point mutations within the conserved regulatory sites HF-1a and HF-1b significantly cripple ventricular muscle specificity, while mutation of the single E-box site was without effect, suggesting that ventricular muscle-specific expression occurs through an E-box-independent pathway. This study provides direct evidence that the cis regulatory sequences in the cardiac/slow twitch MLC-2 gene which confer cardiac and skeletal muscle-specific expression can be clearly segregated, suggesting that distinct regulatory programs may have evolved to control the tissue-specific expression of this single contractile protein gene in cardiac and skeletal muscle.

Non-Q-wave myocardial infarction: incidence, pathophysiology, and clinical course compared with Q-wave infarction.

Acute myocardial infarction can be stratified into electrocardiographic subsets based on the presence or absence of new Q waves. This stratification has important clinical and prognostic utility. Coronary angiography during acute non-Q-wave compared with Q-wave infarction shows much higher infarct-related artery patency rates (about 75 vs. 10%) and increased coronary collateralization. Culprit legion morphology in non-Q-wave infarction commonly is complex. The frequent demonstration of recurrent clinical ischemia, or residual thallium-201 uptake abnormalities, and metabolic activity on persistent emission tomography suggest the presence of viable myocardium in the distribution of the non-Q-wave infarction usually represents 20-25% of acute myocardial infarctions. The University of California San Diego Collaborative Postinfarction Database and other large studies have found non-Q-wave infarctions to be more common in patients with a history of previous infarction and congestive heart failure, although their mortality during acute hospitalization is lower. However, in long-term followup to one year and beyond, non-Q-wave infarct mortality rates equal those of Q-wave infarction. Patients at low early and late risk of mortality include those with a first infarction who are under age 70, whereas patients with evidence of residual ischemia postinfarction are at increased risk of events. It is interesting to speculate that the settings of unstable angina pectoris, non-Q-wave infarction, and perhaps the picture after thrombolysis for acute myocardial infarction, have pathophysiologic similarities which may carry implications for future research and therapy.

Demonstration of cross-reactive antibodies to smooth gram-negative bacteria in antiserum to Escherichia coli J5.

We investigated the discrepancy between the broad cross-protection against gram-negative infections afforded by antiserum to Escherichia coli J5 and its apparently narrow cross-reactivity in vitro. Rabbits immunized with J5 bacteria produced antibodies to both the J5 lipopolysaccharide (LPS; titer by ELISA, 1:60,000) and LPS from the Re mutant of Salmonella minnesota (i.e., to the ketodeoxyoctonate [KDO] and lipid A determinants; titer, 1:3,200). In highly diluted antiserum, titers of antibody to J5 LPS were reduced by 28%-41% after adsorption with seven strains of smooth gram-negative bacteria and by only 4% after adsorption with the Re mutant. Smooth gram-negative bacteria adsorbed virtually all antibody to Re LPS. Therefore, rabbit antiserum to J5 contains type-specific antibodies to core determinants distal to KDO that can obscure highly cross-reactive antibodies to lipid A-KDO in vitro. Cross-reactive antibodies are demonstrable by adsorption with whole bacteria at limiting concentrations of antibody.