Serum but not myocardial TNF-alpha concentration is increased in pacing-induced heart failure in rabbits

In animals and patients with severe heart failure (HF), the serum tumor necrosis factor-alpha (TNF-alpha) concentration is increased. It is, however, still controversial whether or not such increased serum TNF-alpha originates from the heart itself or is of peripheral origin secondary to gastrointestinal congestion and increased endotoxin concentration. We therefore now examined TNF-alpha in serum, myocardium, and liver of sham-operated and HF rabbits. In nine rabbits in which HF was induced by left ventricular (LV) pacing at 400 beats/min for 3 wk, LV end-diastolic diameter was increased and systolic shortening fraction (9.4 +/- 1.0 vs. 28.5 +/- 1.3%, echocardiography, P < 0.05) was reduced. Serum TNF-alpha was higher in HF than in sham-operated rabbits (240 +/- 24 vs. 150 +/- 22 U/ml, WEHI-cell assay, P < 0.05). In the heart, TNF-alpha was located mainly in the vascular endothelium (immunohistochemistry), and TNF-alpha protein (920 +/- 160 vs. 900 +/- 95 U/g) did not differ between groups. In the liver of HF rabbits, hepatocytes expressed TNF-alpha, and TNF-alpha protein was increased compared with sham-operated rabbits (2,390 +/- 310 vs. 1,220 +/- 135 U/g, P < 0.05) and correlated to the number of hepatic leukocytes (r = 0.85) and serum TNF-alpha (r = 0.69). The intestinal endotoxin concentration was 24.5 +/- 1.2 vs. 17.0 +/- 3.1 endotoxin units/g wet wt (P < 0.05) in HF compared with sham-operated rabbits. In this HF model, serum but not myocardial TNF-alpha is increased. The increased serum TNF-alpha originates from peripheral sources.     

Enhanced peripheral chemoreflex function in conscious rabbits with pacing-induced heart failure.

The present study aimed to determine whether peripheral and/or central chemoreflex function is altered in chronic heart failure (CHF) and whether altered chemoreflex function contributes to sympathetic activation in CHF. A rabbit model of pacing-induced CHF was employed. The development of CHF (3-4 wk of pacing) was characterized by an enlarged heart, an attenuated contractility, and an elevated central venous pressure. Renal sympathetic nerve activity (RSNA) and minute volume (MV) of ventilation in response to stimulation of peripheral chemoreceptors by isocapnic/hypoxic gases were measured in the conscious state. It was found that the baseline RSNA at normoxia was higher in CHF rabbits than in sham rabbits (35. 00 +/- 4.03 vs. 20.75 +/- 2.87% of maximum, P < 0.05). Moreover, the magnitudes of changes in RSNA and MV in response to stimulation of the peripheral chemoreceptors and the slopes of RSNA-arterial PO2 and MV-arterial PO2 curves were greater in CHF than in sham rabbits. Inhibition of the peripheral chemoreceptors by inhalation of 100% O2 decreased RSNA in CHF but not in sham rabbits. The central chemoreflex function, as evaluated by the responses of RSNA and MV to hyperoxic/hypercapnic gases, was not different between sham and CHF rabbits. These data suggest that an enhancement of the peripheral chemoreflex occurs in the rabbit model of pacing-induced CHF and that the enhanced peripheral chemoreflex function contributes to the sympathetic activation in the CHF state.     

Protein synthesis is increased in heart failure induced by low dose adriamycin in rabbits

Congestive heart failure was induced in rabbits by a chronic treatment with a low dose of adriamycin (0.75 mg/kg intravenously 3 times per week for 11 weeks). Twenty-four to 48 h after the last injection, adriamycin-treated rabbits had a three-fold increase in plasma norepinephrine, a seven-fold increase in plasma epinephrine, a 19 +/- 8% increase in heart rate, and a 54 +/- 10% decrease in the total tension generated by their isolated papillary muscles, when compared with normal age-matched controls. This demonstrated the occurrence of the cardiomyopathy and heart failure. The effect of adriamycin on myocardial and diaphragmatic protein synthesis was examined in vivo after a 1-h infusion with [3H]leucine and in vitro after a 2-h incubation of right ventricular papillary muscle with [3H]leucine. The rate of in vivo [3H]leucine incorporation into total protein was increased in the heart of the adriamycin-treated rabbits. The increases were 60 +/- 16% in the left ventricle, 49 +/- 18% in the septum, 32 +/- 18% in the right ventricle, and 66 +/- 16% in the atria. A similar increase was observed when measuring the rate of [3H]leucine incorporation into myosin, a myofibrillar protein, and when the rate of [3H]leucine incorporation into total protein was measured in vitro in papillary muscle. In contrast, the rate of [3H]leucine incorporation into total protein of the diaphragm was not significantly changed.(ABSTRACT TRUNCATED AT 250 WORDS)     

Akt signaling pathway in pacing-induced heart failure

Marked changes in energy substrate utilization occur during the progression of congestive heart failure (CHF) where fatty acid utilization, as the primary source of cardiac energy, is severely diminished, oxidative phosphorylation is down-regulated, and glucose uptake and utilization increase. Neither the signaling events or the molecular basis for the shift in substrate utilization have yet been elucidated. This study was designed to examine in the canine model of paced-induced CHF, the potential role of the Akt pathway in signaling the metabolic transitions central to progression to heart failure. Myocardial Akt levels were elevated in early heart failure (after 1–2 weeks of pacing) accompanied by increased levels of oxidative stress, cytokine tumor necrosis factor- (TNF-) and free fatty acid accumulation, reduced activity levels of mitochondrial respiratory complexes III and V and apoptosis initiation. At severe heart failure (3–4 weeks of pacing), there was significant further increase in myocardial apoptosis, with pronounced decline in myocardial Akt kinase activity. At this later stage, there were no further changes in free fatty acid accumulation, complex V activity or in oxidative stress levels indicating that these changes primarily occurred in the earlier stage of evolving heart failure. In contrast, during severe heart failure, both the reduction in complex III activity and increase in TNF- level became more pronounced. Our data provide critical support for the hypothesis that the Akt signaling pathway is a contributory element in the early signaling events leading to the progression of pacing-induced heart failure, accompanying the shift in substrate utilization. (Mol Cell Biochem 268: 103–110, 2005)     

Phosphodiesterase 3 activity is reduced in dog lung following pacing-induced heart failure

We hypothesized that decreases in expression and/or activity of cAMP-specific phosphodiesterases (PDE) contribute to protective adaptations observed in lung after heart failure. In this study, we compared PDE activity in lung parenchyma isolated from control dogs and those paced to heart failure by assaying cyclic nucleotide hydrolysis in fractions of homogenate supernatant eluted from DEAE-Trisacryl columns. Cyclic nucleotide hydrolysis due to PDE3, PDE4, and PDE5 isoforms was predominant in both control and paced groups. The ratio of PDE3 activity to total cAMP PDE activity was decreased in the paced group compared with control (P < 0.05), whereas PDE4 or PDE5 activity ratios were not different between the two groups. With the use of RT-PCR, message expression for PDE3A or PDE3B did not differ between the two groups. Cilostamide, a selective PDE3 inhibitor, and forskolin, a nonspecific agonist for adenylyl cyclase, both inhibited thapsigargin-induced increases in endothelial permeability in control lung. We conclude that PDE3 activity, but not mRNA expression, is reduced in lung from dogs paced to heart failure, a change that could contribute to heart failure-induced attenuation of the lung endothelial permeability response to injury.     

Asymmetrical myocardial expression of natriuretic peptides in pacing-induced heart failure

High-frequency pacing of the left ventricle (LV) free wall causes a dyssynchronous pattern of contraction that leads to progressive heart failure (HF) with pronounced differences in regional contractility. Aim of this study was to evaluate possible changes in brain natriuretic peptide (BNP) and C-type natriuretic peptide (CNP) mRNA expression in the anterior/anterior lateral region (pacing site, PS) as compared to the infero-septal region (opposite site, OS) and to explore possible association between the contractiling pattern and biomarker expression. Cardiac tissue was collected from minipigs with pacing-induced HF (n = 8) and without (control, n = 6). The samples were selectively harvested from the anterior left ventricular (LV) wall, PS, and from an area remote to the pacing-site, OS. BNP and CNP mRNA expression was evaluated by semi-quantitative polymerase chain reaction (PCR). A significant difference in BNP expression was found in the PS between HF animals and controls (BNP/GAPDH: 0.65 ± 0.11 vs. 0.35 ± 0.04, p = 0.02), but not in the OS (BNP/GAPDH: 0.36 ± 0.05, ns vs. controls). CNP expression was not different compared to controls, although higher levels were observed in the PS and in the OS with respect to the controls (CNP/GAPDH: controls 0.089 ± 0.036, PS 0.289 ± 0.23, OS 0.54 ± 0.16). This finding was in tune with an increase of CNP tissue concentration (controls: 0.69 ± 0.13; PS = 1.56 ± 0.19; OS = 1.70 ± 0.42 pg/mg protein; p = 0.039 controls vs. OS). Higher BNP mRNA expression in the PS is consistent with a reduction in contractile function in this region, while higher CNP mRNA expression in the OS suggests the presence of concomitant endothelial dysfunction in the remote region.     

Implications of QRS duration in dogs with pacing-induced heart failure

The objective of this study was to find out the implication of QRS duration in dogs with rapid pacing-induced heart failure. Sixteen Beagle dogs were implanted with transvenous cardiac pacemakers and underwent rapid right ventricular pacing for 3 weeks at 260 bpm to induce heart failure. Dogs were divided into two groups according to the QRS duration: 9 with normal QRS duration (<100 ms) and 7 with prolonged QRS duration (≥100 ms). Cardiac systolic function and size was analyzed by real time 3-dimensional echocardiography and left ventricular dyssynchrony was assessed by speckle tracking strain imaging. Congestive heart failure developed 3 weeks after rapid right ventricular pacing. Dogs with prolonged QRS duration showed more extensive radial strain and circumferential strain dyssynchrony than dogs with normal QRS duration. At the end of 4-week recovery, greater improvement of left ventricular ejection fraction and left ventricular end-systolic volume was detected in dogs with normal QRS duration. The findings suggested that left ventricular dyssynchrony, indicated by a prolonged QRS duration, predicted an unsatisfying recovery in dogs with rapid pacing-induced heart failure. QRS duration had the potential to be a prognostic indicator for dogs with heart failure.     

Alveolar fluid reabsorption is increased in rats with compensated heart failure

Alveolar fluid reabsorption (AFR) is important in keeping the air spaces free of edema. This process is accomplished via active transport of Na(+) across the alveolo-capillary barrier mostly by apical Na(+) channels and basolateral Na(+)-K(+)-ATPases. Recently, we have reported that acute elevation of left atrial pressures is associated with decreased AFR in isolated rat lungs. However, the effect of chronic elevation of pulmonary capillary pressure, such as seen in patients with congestive heart failure (CHF), on AFR is unknown. CHF was induced by creating an aorto-caval fistula (ACF) in Sprague-Dawley male rats. Seven days after the placement of the fistula, AFR was studied in the isolated perfused rat lung model. AFR in control rats was 0.49 +/- 0.02 ml/h (all values are means +/- SE) and increased by approximately 40% (0.69 +/- 0.03 ml/h) in rats with chronic CHF (P < 0.001). The albumin flux from the pulmonary circulation into the air spaces did not increase in the experimental groups, indicating that lung permeability for large solutes was not increased. Na(+)-K(+)-ATPase activity and protein abundance at the plasma membrane of distal alveolar epithelial tissue were significantly increased in CHF rats compared with controls. These changes were associated with increased plasma norepinephrine levels in CHF rats compared with controls. We provide evidence that in a rat model of chronic compensated CHF, AFR is increased, possibly due to increased endogenous norepinephrine upregulating active sodium transport and protecting against alveolar flooding.     

Renin-angiotensin system inhibition on noradrenergic nerve terminal function in pacing-induced heart failure

Chronic angiotensin-converting enzyme (ACE) inhibition has been shown to improve cardiac sympathetic nerve terminal function in heart failure. To determine whether similar effects could be produced by angiotensin II AT(1) receptor blockade, we administered the ACE inhibitor quinapril, angiotensin II AT(1) receptor blocker losartan, or both agents together, to rabbits with pacing-induced heart failure. Chronic rapid pacing produced left ventricular dilation and decline of fractional shortening, increased plasma norepinephrine (NE), and caused reductions of myocardial NE uptake activity, NE histofluorescence profile, and tyrosine hydroxylase immunostained profile. Administration of quinapril or losartan retarded the progression of left ventricular dysfunction and attenuated cardiac sympathetic nerve terminal abnormalities in heart failure. Quinapril and losartan together produced greater effects than either agent alone. The effect of renin-angiotensin system inhibition on improvement of left ventricular function and remodeling, however, was not sustained. Our results suggest that the effects of ACE inhibitors are mediated via the reduction of angiotensin II and that angiotensin II plays a pivotal role in modulating cardiac sympathetic nerve terminal function during development of heart failure. The combined effect of ACE inhibition and angiotensin II AT(1) receptor blockade on cardiac sympathetic nerve terminal dysfunction may contribute to the beneficial effects on cardiac function in heart failure.     

Effect of PDE5 inhibition on coronary hemodynamics in pacing-induced heart failure

Inhibition of phosphodiesterase type 5 (PDE5) can relax systemic and coronary vessels by causing accumulation of cGMP. Both the endothelial dysfunction with decreased nitric oxide production and increased natriuretic peptide levels in congestive heart failure (CHF) have the potential to alter cGMP production, thereby influencing the response to PDE5 inhibition. Consequently, this study examined the effects of PDE5 inhibition with sildenafil in dogs with CHF produced by rapid ventricular pacing. CHF resulted in decreases of left ventricular (LV) systolic pressure, coronary blood flow, and the maximal first time derivative of LV pressure (LV dP/dt(max)) at rest and during treadmill exercise compared with normal, whereas resting LV end-diastolic pressure increased from 10 +/- 1.4 to 23 +/- 1.4 mmHg. Sildenafil (2 and 10 mg/kg per os) caused a 5- to 6-mmHg decrease of aortic pressure (P < 0.05), with no change of heart rate, LV systolic pressure, or LV dP/dt(max). Sildenafil caused no change in coronary flow or myocardial oxygen consumption in animals with CHF at rest or during exercise. In contrast to findings in normal animals, sildenafil did not augment endothelium-dependent coronary vasodilation in response to acetylcholine in animals with CHF. Furthermore, Western blotting showed decreased PDE5 protein expression in myocardium from failing hearts. These findings demonstrate that PDE5 contributes little to regulation of coronary hemodynamics in CHF.     

Hemodynamic and norepinephrine responses to pacing-induced heart failure in conscious sinoaortic-denervated dogs

Brändle, Marian, Kaushik P. Patel, Wei Wang, andIrving H. Zucker. Hemodynamic and norepinephrine responses topacing-induced heart failure in conscious sinoaortic-denervated dogs.J. Appl. Physiol. 81(4):1855-1862, 1996.The present study was undertaken to determinethe effects of chronic sinoaortic (baroreceptor) denervation (SAD) on the hemodynamic and sympathetic alterations thatoccur in the pacing-induced model of congestive heart failure. Twogroups of dogs were examined: intact(n = 9) and SAD(n = 9). Both groups of dogs werestudied in the control (prepace) state and each week after theinitiation of ventricular pacing at 250 beats/min. After the pacemakerwas turned off, hemodynamic and plasma norepinephrine levels returnedtoward control levels in the prepaced state and after 1 and 2 wk ofpacing. However, by 3 wk all hemodynamic and norepinephrine levelsremained relatively constant over the 10-min observation period withthe pacemaker off. With the pacemaker off, left ventricularend-diastolic pressure went from 2.7 ± 1.4 (SE) mmHg during theprepace state to 23.2 ± 2.9 mmHg in the heart failure state inintact dogs (P < 0.01). Leftventricular end-diastolic pressure increased to 27.1 ± 2.2 mmHgfrom a control level of 4.2 ± 1.9 mmHg in SAD dogs(P < 0.0003). Mean arterial pressuresignificantly decreased in intact and SAD dogs. Resting heart rate wassignificantly higher in SAD dogs and increased to 135.8 ± 8.9 beats/min in intact dogs and 136.1 ± 6.5 beats/min in SAD dogs.There were no significant differences in the hemodynamic parametersbetween intact and SAD dogs after pacing. Plasma norepinephrine wassignificantly lower in intact than in SAD dogs before pacing (197.7 ± 21.6 vs. 320.6 ± 26.6 pg/ml;P < 0.005). In the heart failurestate, plasma norepinephrine increased significantly in both intact(598.3 ± 44.2 pg/ml) and SAD (644.0 ± 64.6 pg/ml) groups. Therewere no differences in the severity or the magnitude of the developedheart failure state in SAD vs. intact dogs. We conclude from these datathat the arterial baroreflex is not the sole mechanism for the increasein sympathetic drive in heart failure.

     

Alterations in cardiac adrenergic terminal function and beta-adrenoceptor density in pacing-induced heart failure

Congestive heart failure is associated with cardiac adrenergic nerve terminal changes and beta-adrenoceptor density downregulation. To study the temporal sequence of these changes, we performed studies in rabbits at 2, 4, and 8 wk of cardiac pacing (360 beats/min) and at 1, 2, and 4 wk after cessation of pacing. Rapid pacing produced left ventricular (LV) dysfunction and an increase in plasma norepinephrine (NE) in 1-2 wk. At week 2, NE uptake activity, NE uptake-1 density, and adenylyl cyclase responses to isoproterenol, 5'-guanylyl imidodiphosphate [Gpp(NH)p], and forskolin reduced. However, immunostained tyrosine hydroxylase profile, beta-adrenoceptor density, and NE histofluorescence did not reduce until 4-8 wk of pacing. After cessation of cardiac pacing, LV function normalized quickly, followed by return of tyrosine hydroxylase and NE profiles in 1 wk and adenylyl cyclase responses to agonists and NE uptake activity in 2 wk. Myocardial beta-adrenoceptor density returned to normal by 4 wk after cessation of pacing. Our results suggest that there is no permanent structural neuronal damage in the myocardium within the first 8 wk of rapid cardiac pacing. Abnormal myocardial NE reuptake mechanism may play an important pathophysiological role in heart failure.     

EMT phenotype is induced by increased Src kinase activity via Src-mediated caspase-8 phosphorylation

Caspase-8 governs multiple cell responses to the microenvironmental cues. However, its integration of "death-life" signalings remains elusive. In our study, the role of caspase-8-Src is well-established as a promoter for migration or metastasis in Casp8(+)Src(+) A549/H226 cells in vivo and in vitro. In particular for nude mice models, mice implanted with Casp8(+)Src(+) A459/H226 cells remarkably increased spontaneous tumor metastatic burden with a significant survival disadvantage. Additionally, we detect that Src-mediated caspase-8 phosphorylation stimulates Src phosphorylation at Tyr-416 via the linkage of Src SH2 domain with phosph-Tyr-380 site of caspase-8. In turn, activated Src can efficiently induce epithelial-mesenchymal transition (EMT) phenotypic features to promote tumor cells metastasis. Surprisingly, RXDLL motif deletion in the DEDa of caspase-8 attenuates tumor cell migration or metastasis via impairing the recruitment of caspase-8 into the cellular periphery where activated Src is subject to caspase-8 phosphorylation. Together, a simple model is that the peripherization of caspase-8 is well-poised to facilitate Src-mediated caspase-8 phosphrylation at Tyr-380, then binding of phospho-Tyr380 of caspase-8 to Src SH2 domain may maintain Src in an active conformation to induce EMT phenotype, a key step toward cancer metastasis.     

Cardiac mitochondria in heart failure: normal cardiolipin profile and increased threonine phosphorylation of complex IV

Mitochondrial dysfunction is a major contributor in heart failure (HF). We investigated whether the decrease in respirasome organization reported by us previously in cardiac mitochondria in HF is due to changes in the phospholipids of the mitochondrial inner membrane or modifications of the subunits of the electron transport chain (ETC) complexes. The contents of the main phospholipid species, including cardiolipin, as well as the molecular species of cardiolipin were unchanged in cardiac mitochondria in HF. Oxidized cardiolipin molecular species were not observed. In heart mitochondria isolated from HF, complex IV not incorporated into respirasomes exhibits increased threonine phosphorylation. Since HF is associated with increased adrenergic drive to cardiomyocytes, this increased protein phosphorylation might be explained by the involvement of cAMP-activated protein kinase. Does the preservation of cAMP-induced phosphorylation changes of mitochondrial proteins or the addition of exogenous cAMP have similar effects on oxidative phosphorylation? The usage of phosphatase inhibitors revealed a specific decrease in complex I-supported respiration with glutamate. In saponin-permeabilized cardiac fibers, pre-incubation with cAMP decreases oxidative phosphorylation due to a defect localized at complex IV of the ETC inter alia. We propose that phosphorylation of specific complex IV subunits decreases oxidative phosphorylation either by limiting the incorporation of complex IV in supercomplexes or by decreasing supercomplex stability.     

Ca2+ clock malfunction in a canine model of pacing-induced heart failure

The mechanisms of sinoatrial node (SAN) dysfunction in heart failure (HF) remain unclear. We hypothesized that impaired rhythmic spontaneous sarcoplasmic reticulum Ca(2+) release (Ca(2+) clock) plays an important role in SAN dysfunction in HF. HF was induced in canine hearts by rapid ventricular pacing. The location of pacemaking sites was determined in vivo using computerized electrical mapping in acute open-chest preparations (normal, n = 3; and HF, n = 4). Isoproterenol (Iso, 0.2 μg·kg(-1)·min(-1)) infusion increased heart rate and shifted the pacemaking site to the superior SAN in all normal hearts. However, in failing hearts, Iso did not induce superior shift of the pacemaking site despite heart rate acceleration. Simultaneous optical recording of intracellular Ca(2+) and membrane potential was performed in Langendorff-perfused isolated right atrium (RA) preparations from normal (n = 7) and failing hearts (n = 6). Iso increased sinus rate, enhanced late diastolic Ca(2+) elevation (LDCAE), and shifted the pacemaking sites to the superior SAN in all normal but in none of the HF RAs. Caffeine (2 ml, 20 mmol/l) caused LDCAE and increased heart rate in four normal RAs but in none of the three HF RAs. Iso induced ectopic beats from lower crista terminalis in five of six HF RAs. These ectopic beats were suppressed by ZD-7288, a specific pacemaker current (I(f)) blocker. We conclude that HF results in the suppression of Ca(2+) clock, resulting in the unresponsiveness of superior SAN to Iso and caffeine. HF also increases the ectopic pacemaking activity by activating the I(f) at the latent pacemaking sites in lower crista terminalis.     

Protein kinase C-mediated desmin phosphorylation is related to myofibril disarray in cardiomyopathic hamster heart

The cardiomyopathic (CM) Syrian golden hamster (strain UM-X7.1) exhibits a hereditary cardiomyopathy, which causes premature death resulting from congestive heart failure. The CM animals show extensive cardiac myofibril disarray and myocardial calcium overload. The present study has been undertaken to examine the role of desmin phosphorylation in myofibril disarray observed in CM hearts. The data from skinned myofibril protein phosphorylation assays have shown that desmin can be phosphorylated by protein kinase C (PKC). There is no significant difference in the content of desmin between CM and control hamster hearts. However, the desmin from CM hearts has a higher phosphorylation level than that of the normal hearts. Furthermore, we have examined the distribution of desmin and myofibril organization with immunofluorescent microscopy and immunogold electron microscopy in cultured cardiac myocytes after treatment with the PKC-activating phorbol ester, 12-O-tetradecanylphorbol-13-acetate (TPA). When the cultured normal hamster cardiac cells are treated with TPA, desmin filaments are disassembled and the myofibrils become disarrayed. The myofibril disarray closely mimics that observed in untreated CM cultures. These results suggest that disassembly of desmin filaments, which could be caused by PKC-mediated phosphorylation, may be a factor in myofibril disarray in cardiomyopathic cells and that the intermediate filament protein, desmin, plays an important role in maintaining myofibril alignment in cardiac cells.     

Remodeling of lung interstitium but not resistance vessels in canine pacing-induced heart failure.

We previously showed that pacing-induced heart failure in dogs results in an enhancement of pulmonary vascular reactivity. In the present study we hypothesized that enhanced matrix deposition and structural remodeling of lung resistance microvessels would underlie these functional changes. Using biochemical measures, we found no difference in the normalized lung content of hyaluronan, uronic acid, and collagen between control dogs and dogs paced for 1 mo, although lung dry weight and noncollagen protein content increased significantly in the paced group (P < 0.05). From separate Formalin-fixed lung lobes, 5-microm frozen sections were prepared and stained with Masson's trichrome, and vascular structure was evaluated using standard morphometric techniques. When perivascular fluid cuffs were excluded from the measure of wall thickness, collagen and media volume fractions in any size range did not differ between paced and control groups. Similarly, in the paced group, medial thickness in <400-microm arterial or venular microvessels did not vary significantly from that in the controls. In contrast, the relationship of interstitial fluid pressure to lung water was significantly shifted to the right in the paced group, such that normal tissue pressures were observed, despite the increased water content. We conclude that although 1 mo of pacing-induced heart failure results in altered interstitial function, the attendant pulmonary hypertension and/or hormonal responses are insufficient to induce medial hypertrophy or other remodeling of the extra-alveolar microvasculature.     

Impact of enalapril therapy on in vitro coronary artery responsiveness in pacing-induced heart failure.

In vitro coronary artery responsiveness to angiotensin I, angiotensin II, noradrenaline, phenylephrine, BHT 920, and potassium chloride together with functional relaxation to acetylcholine was investigated in dogs with pacing-induced heart failure treated with enalapril (oral administration of 10 mg.day-1) for a mean duration of 26 days. Although maximal responses generated to both angiotensin I and angiotensin II were unaltered in the enalapril-treated group, angiotensin II became more potent following enalapril treatment: the EC50 for angiotensin II following placebo treatment was 2.4 (0.6-5.8; 95% confidence limits) nM and following enalapril treatment was 0.03 (0.007-0.1; 95% confidence limits) nM. In addition to the above changes, coronary artery rings from dogs treated with enalapril developed significantly less tension to noradrenaline, phenylephrine, and BHT 920. In contrast, responses to potassium chloride were unaltered following enalapril treatment. However, the relaxation to acetylcholine was enhanced from 38.9 +/- 3.0 to 50.4 +/- 3.5% (placebo versus enalapril, p < 0.05). These findings indicate that enalapril may possess alpha-blocking properties and enhance the relaxation response to acetylcholine through an endothelial-dependent mechanism in addition to inhibiting converting enzyme.     

Myosin I phosphorylation is increased by chemotactic stimulation

Directed cell migration occurs in response to extracellular cues. Following stimulation of a cell with chemoattractant, a significant rearrangement of the actin cytoskeleton is mediated by intracellular signaling pathways and results in polarization of the cell and movement via pseudopod extension. Amoeboid myosin Is play a critical role in regulating pseudopod formation in Dictyostelium, and their activity is activated by heavy chain phosphorylation. The effect of chemotactic stimulation on the in vivo phosphorylation level of a Dictyostelium myosin I, myoB, was tested. The myoB heavy chain is phosphorylated in vivo on serine 322 (the myosin TEDS rule phosphorylation site) in chemotactically competent cells. The level of myoB phosphorylation increases following stimulation of starving cells with the chemoattractant cAMP. A 3-fold peak increase in the level of phosphorylation is observed at 60 s following stimulation, a time at which the Dictyostelium cell actively extends pseudopodia. These findings suggest that chemotactic stimulation results in increased myoB activity via heavy chain phosphorylation and contributes to the global extension of pseudopodia that occurs prior to polarization and directed motility.     

Dynamic changes in conduction velocity and gap junction properties during development of pacing-induced heart failure

End-stage heart failure (HF) is characterized by changes in conduction velocity (CV) that predispose to arrhythmias. Here, we investigate the time course of conduction changes with respect to alterations in connexin 43 (Cx43) properties and mechanical function during the development of HF. We perform high-resolution optical mapping in arterially perfused myocardial preparations from dogs subjected to 0, 3, 7, 14, and 21 days of rapid pacing to produce variable degrees of remodeling. CV is compared with an index of mechanical function [left ventricular end-diastolic pressure (LVEDP)] and with dynamic changes in the expression, distribution, and phosphorylation of Cx43. In contrast to repolarization, CV was preserved during early stages of remodeling (3 and 7 days) and significantly reduced at later stages, which were associated with marked increases in LVEDP. Measurements of differentially phosphorylated Cx43 isoforms revealed early, sustained downregulation of pan-Cx43 that preceded changes in CV and LVEDP, a gradual rise in a dephosphorylated Cx43 isoform to over twofold baseline levels in end-stage HF, and a late abrupt increase in pan-Cx43, but not dephosphorylated Cx43, lateralization. These data demonstrate that 1) CV slowing occurs only at advanced stages of remodeling, 2) total reduction of pan-Cx43 is an early event that precedes mechanical dysfunction and CV slowing, 3) changes in Cx43 phosphorylation are more closely associated with the onset of HF, and 4) Cx43 lateralization is a late event that coincides with marked CV reduction. These data reveal a novel paradigm of remodeling based on the timing of conduction abnormalities relative to changes in Cx43 isoforms and mechanical dysfunction.