Changes in the pro-inflammatory cytokine production and peritoneal macrophage function in rats with chronic heart failure

Chronic heart failure (CHF) is a state of immune activation, and pro-inflammatory cytokines play an important role in its development and progression. Macrophages (Mphis), when activated, are the main source of pro-inflammatory cytokines. We measured interleukin-6 (IL-6), interleukin (IL-1beta) and tumor necrosis factor - alpha (TNF-alpha) production after lipopolysaccharide (LPS)-stimulation, as well as peritoneal Mphis migration, phagocytic capacity, chemotaxis index, and hydrogen peroxide production, in an attempt to clarify the role of this cell in an animal model of CHF. Ligature of the left coronary artery or sham operation was performed in adult Wistar rats. After 12 weeks, resident and total cell number, phagocytic capacity, chemotaxis index, and hydrogen peroxide production in Mphis were significantly higher in CHF than in control rats. The production of IL-6 and TNF- alpha was similarly significantly enhanced in CHF as compared with controls. Mphis obtained from CHF rats were more responsive to LPS, suggesting the existence, in vivo, of possible factor(s) modulating the production of pro-inflammatory cytokines. The results demonstrated that there is modification of peritoneal Mphis function along CHF development, possibly contributing to the pathophysiological process in the establishment of CHF.     

Proteasome inhibition improves diaphragm function in congestive heart failure rats

In congestive heart failure (CHF), diaphragm weakness is known to occur and is associated with myosin loss and activation of the ubiquitin-proteasome pathway. The effect of modulating proteasome activity on myosin loss and diaphragm function is unknown. The present study investigated the effect of in vivo proteasome inhibition on myosin loss and diaphragm function in CHF rats. Coronary artery ligation was used as an animal model for CHF. Sham-operated rats served as controls. Animals were treated with the proteasome inhibitor bortezomib (intravenously) or received saline (0.9%) injections. Force generating capacity, cross-bridge cycling kinetics, and myosin content were measured in diaphragm single fibers. Proteasome activity, caspase-3 activity, and MuRF-1 and MAFbx mRNA levels were determined in diaphragm homogenates. Proteasome activities in the diaphragm were significantly reduced by bortezomib. Bortezomib treatment significantly improved diaphragm single fiber force generating capacity (approximately 30-40%) and cross-bridge cycling kinetics (approximately 20%) in CHF. Myosin content was approximately 30% higher in diaphragm fibers from bortezomib-treated CHF rats than saline. Caspase-3 activity was decreased in diaphragm homogenates from bortezomib-treated rats. CHF increased MuRF-1 and MAFbx mRNA expression in the diaphragm, and bortezomib treatment diminished this rise. The present study demonstrates that treatment with a clinically used proteasome inhibitor improves diaphragm function by restoring myosin content in CHF.     

Rosiglitazone treatment restores renal responsiveness to atrial natriuretic peptide in rats with congestive heart failure

The thiazolidinedione (TZD) class of Peroxisome proliferator‐activated receptor gamma agonists has restricted clinical use for diabetes mellitus due to fluid retention and potential cardiovascular risks. These side effects are attributed in part to direct salt‐retaining effect of TZDs at the renal collecting duct. A recent study from our group revealed that prolonged rosiglitazone (RGZ) treatment caused no Na+/H₂O retention or up‐regulation of Na⁺ transport‐linked channels/transporters in experimental congestive heart failure (CHF) induced by surgical aorto‐caval fistula (ACF). The present study examines the effects of RGZ on renal and cardiac responses to atrial natriuretic peptide (ANP), Acetylcholine (Ach) and S‐Nitroso‐N‐acetylpenicillamine (SNAP‐NO donor). Furthermore, we assessed the impact of RGZ on gene expression related to the ANP signalling pathway in animals with ACF. Rats subjected to ACF (or sham) were treated with either RGZ (30 mg/kg/day) or vehicle for 4 weeks. Cardiac chambers pressures and volumes were assessed invasively via Miller catheter. Kidney excretory and renal hemodynamic in response to ANP, Ach and SNAP were examined. Renal clearance along with cyclic guanosine monophosphate (cGMP), gene expression of renal CHF‐related genes and ANP signalling in the kidney were determined. RGZ‐treated CHF rats exhibited significant improvement in the natriuretic responses to ANP infusion. This ‘sensitization’ to ANP was not associated with increases in neither urinary cGMP nor in vitro cGMP production. However, RGZ caused down‐regulation of several genes in the renal cortex (Ace, Nos3 and Npr1) and up‐regulation of ACE2, Agtrla, Mme and Cftr along down‐regulation of Avpr2, Npr1,2, Nos3 and Pde3 in the medulla. In conclusion, CHF+RGZ rats exhibited significant enhancement in the natriuretic responses to ANP infusion, which are known to be blunted in CHF. This ‘sensitization’ to ANP is independent of cGMP signalling, yet may involve post‐cGMP signalling target genes such as ACE2, CFTR and V2 receptor. The possibility that TZD treatment in uncomplicated CHF may be less detrimental than thought before deserves additional investigations.     

Metformin improves cardiac function in a nondiabetic rat model of post-MI heart failure

Metformin is the first choice drug for the treatment of patients with diabetes, but its use is debated in patients with advanced cardiorenal disease. Epidemiological data suggest that metformin may reduce cardiac events, in patients both with and without heart failure. Experimental evidence suggests that metformin reduces cardiac ischemia-reperfusion injury. It is unknown whether metformin improves cardiac function (remodeling) in a long-term post-MI remodeling model. We therefore studied male, nondiabetic, Sprague-Dawley rats that were subjected to either myocardial infarction (MI) or sham operation. Animals were randomly allocated to treatment with normal water or metformin-containing water (250 mg·kg(-1)·day(-1)). At baseline, 6 wk, and 12 wk, metabolic parameters were analyzed and oral glucose tolerance tests (OGTT) were performed. Echocardiography and hemodynamic parameters were assessed 12 wk after MI. In the MI model, infarct size was significantly smaller after 12-wk metformin treatment (29.6 ± 3.2 vs. 38.0 ± 2.2%, P < 0.05). Moreover, metformin resulted in less left ventricular dilatation (6.0 ± 0.4 vs. 7.6 ± 0.6 mm, P < 0.05) and preservation of left ventricular ejection fraction (65.8 ± 3.7% vs. 48.6 ± 5.6%, P < 0.05) compared with MI control. The improved cardiac function was associated with decreased atrial natriuretic peptide mRNA levels in the metformin-treated group (50% reduction compared with MI, P < 0.05). Insulin resistance did not occur during cardiac remodeling (as indicated by normal OGTT) and fasting glucose levels and the pattern of the OGTT were not affected by metformin. Molecular analyses suggested that altered AMP kinase phosphorylation status and low insulin levels mediate the salutary effects of metformin. Altogether our results indicate that metformin may have potential to attenuate heart failure development after myocardial infarction, in the absence of diabetes and independent of systemic glucose levels.     

Temperature-dependent skeletal muscle dysfunction in rats with congestive heart failure.

Abnormalities in the excitation-contraction coupling of slow-twitch muscle seem to explain the slowing and increased fatigue observed in congestive heart failure (CHF). However, it is not known which elements of the excitation-contraction coupling might be affected. We hypothesize that the temperature sensitivity of contractile properties of the soleus muscle might be altered in CHF possibly because of alterations of the temperature sensitivity of intracellular Ca(2+) handling. We electrically stimulated the in situ soleus muscle of anesthetised rats that had 6-wk postinfarction CHF using 1 and 50 Hz and using a fatigue protocol (5-Hz stimulation for 30 min) at 35, 37, and 40 degrees C. Ca(2+) uptake and release were measured in sarcoplasmic reticulum vesicles at various temperatures. Contraction and relaxation rates of the soleus muscle were slower in CHF than in sham at 35 degrees C, but the difference was almost absent at 40 degrees C. The fatigue protocol revealed that force development was more temperature sensitive in CHF, whereas contraction and relaxation rates were less temperature sensitive in CHF than in sham. The Ca(2+) uptake and release rates did not correlate to the difference between CHF and sham regarding contractile properties or temperature sensitivity. In conclusion, the discrepant results regarding altered temperature sensitivity of contraction and relaxation rates in the soleus muscle of CHF rats compared with Ca(2+) release and uptake rates in vesicles indicate that the molecular cause of slow-twitch muscle dysfunction in CHF is not linked to the intracellular Ca(2+) cycling.     

A new model of congestive heart failure in rats

Current rodent models of ischemia/infarct or pressure-volume overload are not fully representative of human heart failure. We developed a new model of congestive heart failure (CHF) with both ischemic and stress injuries combined with fibrosis in the remote myocardium. Sprague-Dawley male rats were used. Ascending aortic banding (Ab) was performed to induce hypertrophy. Two months post-Ab, ischemia-reperfusion (I/R) injury was induced by ligating the left anterior descending (LAD) artery for 30 min. Permanent LAD ligation served as positive controls. A debanding (DeAb) procedure was performed after Ab or Ab + I/R to restore left ventricular (LV) loading properties. Cardiac function was assessed by echocardiography and in vivo hemodynamic analysis. Myocardial infarction (MI) size and myocardial fibrosis were assessed. LV hypertrophy was observed 4 mo post-Ab; however, systolic function was preserved. LV hypertrophy regressed within 1 mo after DeAb. I/R for 2 mo induced a small to moderate MI with mild impairment of LV function. Permanent LAD ligation for 2 mo induced large MI and significant cardiac dysfunction. Ab for 2 mo followed by I/R for 2 mo (Ab + I/R) resulted in moderate MI with significantly reduced ejection fraction (EF). DeAb post Ab + I/R to reduce afterload could not restore cardiac function. Perivascular fibrosis in remote myocardium after Ab + I/R + DeAb was associated with decreased cardiac function. We conclude that Ab plus I/R injury with aortic DeAb represents a novel model of CHF with increased fibrosis in remote myocardium. This model will allow the investigation of vascular and fibrotic mechanisms in CHF characterized by low EF, dilated LV, moderate infarction, near-normal aortic diameter, and reperfused coronary arteries.     

Echocardiographic detection of congestive heart failure in postinfarction rats

In studies of congestive heart failure (CHF) treatment, it is essential to select animals with a similar degree of cardiac dysfunction. However, this is difficult to establish without hemodynamic evaluation in rat postinfarction-induced CHF. This study aimed to diagnose CHF in long-term follow-up postinfarction rats using only echocardiographic criteria through a J-tree cluster analysis and Fisher's linear discriminant function. Two sets of sham and infarcted rats were studied. The first was used to perform cluster analysis and the second to prospectively validate the results. Six months after inducing myocardial infarction (MI), rats were subjected to transthoracic echocardiography. Infarct size was measured by histological analysis. Six echocardiographic variables were used in the cluster analysis: left ventricular (LV) systolic dimension, LV diastolic dimension-to-body weight ratio, left atrial diameter-to-body weight ratio, LV posterior wall shortening velocity, E wave, and isovolumetric relaxation time. Cluster analysis joined the rats into one sham and two MI groups. One MI cluster had more severe anatomical and echocardiographic changes and was called MI with heart failure (MI/HF+, n = 24, infarct size: 42.7 ± 5.8%). The other had less severe changes and was called MI without heart failure (MI/HF-, n = 11, infarct size: 32.3 ± 9.9%; P < 0.001 vs. MI/HF+). Three rats with small infarct size (21.6 ± 2.2%) presenting mild cardiac alterations were misallocated in the sham group. Fisher's linear discriminant function was built using these groups and used to prospectively classify additional groups of sham-operated (n = 20) and infarcted rats (n = 57) using the same echocardiographic parameters. The discriminant function therefore detected CHF with 100% specificity and 80% sensitivity considering allocation in MI/HF+ and sham group, and 100% specificity and 58.8% sensitivity considering MI/HF+ and MI/HF- groups, taking into account pathological criteria of CHF diagnosis. Echocardiographic analysis can be used to accurately predict congestive heart failure in postinfarction rats.     

Echocardiographic criteria for detection of postinfarction congestive heart failure in rats.

We evaluated postinfarction myocardial function in rats and determined echocardiographic criteria for congestive heart failure (CHF) using high performance echocardiography. Extensive myocardial infarction (MI) was induced in rats by left coronary occlusion. Sham-operated animals served as controls. Five weeks later, high-frame rate ( approximately 200 Hz), fully digitized, shallow-focus (10-25 mm), two-dimensional, M-mode and Doppler echocardiography was performed. A J-tree cluster analysis was performed using parameters indicative of CHF. Reproducibility was examined. The cluster analysis joined the animals into one Sham and two MI clusters. One of the MI clusters had clinical characteristics of CHF and elevated left ventricular end diastolic pressure. Among the echocardiographic variables, only posterior wall shortening velocity separated the failing and nonfailing MI clusters. We conclude that, by high frame rate echocardiography, it is possible to obtain high- quality recordings in rats. It is feasible to distinguish MI rats with CHF due to myocardial dysfunction from those without failure and to perform longitudinal studies on myocardial function.     

Cheyne-Stokes respiration in congestive heart failure.

Cheyne-Stokes respiration is an abnormal breathing pattern which commonly occurs in patients with decompensated congestive heart failure and neurologic diseases, in whom periods of tachypnea and hyperpnea alternate with periods of apnea. In the majority of these patients, the ventilatory patterns may not be recognized, and the clinical features are generally dominated by the underlying disease process. Cheyne-Stokes respiration may, however, have profound effects on the cardiopulmonary system, causing oxygen desaturation, cardiac arrhythmias, and changes in mental status. Treatment of Cheyne-Stokes respiration in congestive heart failure with supplemental oxygen or nasal continuous positive airway pressure, in addition to conventional therapy, may improve the overall cardiac function and perhaps the patient's prognosis.     

Pacing in congestive heart failure

Despite the major advances in medical drug therapy, heart failure remains a syndrome associated with high mortality and morbidity. Biventricular or left ventricular (LV) short atrioventricular (AV) delay pacing is being tested in congestive heart failure patients with left bundle branch block. The aim is to resynchronise the dyscoordinate LV contraction. A number of studies are underway, but it is clear that while some patients respond remarkably, this is highly variable. Accurate identification of patients likely to benefit will be crucial. The mechanism of benefit is unclear. A greater understanding of the physiological consequences of pacing will be necessary to accurately identify these patients.     

Exercise training improves peripheral chemoreflex function in heart failure rabbits.

An enhancement of peripheral chemoreflex sensitivity contributes to sympathetic hyperactivity in chronic heart failure (CHF) rabbits. The enhanced chemoreflex function in CHF involves augmented carotid body (CB) chemoreceptor activity via upregulation of the angiotensin II (ANG II) type 1 (AT(1))-receptor pathway and downregulation of the neuronal nitric oxide synthase (nNOS)-nitric oxide (NO) pathway in the CB. Here we investigated whether exercise training (EXT) normalizes the enhanced peripheral chemoreflex function in CHF rabbits and possible mechanisms mediating this effect. EXT partially, but not fully, normalized the exaggerated baseline renal sympathetic nerve activity (RSNA) and the response of RSNA to hypoxia in CHF rabbits. EXT also decreased the baseline CB nerve single-fiber discharge (4.9 +/- 0.4 vs. 7.7 +/- 0.4 imp/s at Po(2) = 103 +/- 2.3 Torr) and the response to hypoxia (20.6 +/- 1.1 vs. 36.3 +/- 1.3 imp/s at Po(2) = 41 +/- 2.2 Torr) from CB chemoreceptors in CHF rabbits, which could be reversed by treatment of the CB with ANG II or a nNOS inhibitor. Our results also showed that NO concentration and protein expression of nNOS were increased in the CBs from EXT + CHF rabbits, compared with that in CHF rabbits. On the other hand, elevated ANG II concentration and AT(1)-receptor overexpression of the CBs in CHF state were blunted by EXT. These results indicate that EXT normalizes the CB chemoreflex in CHF by preventing an increase in afferent CB chemoreceptor activity. EXT reverses the alterations in the nNOS-NO and ANG II-AT(1)-receptor pathways in the CB responsible for chemoreceptor sensitization in CHF.     

Statin therapy restores sympathovagal balance in experimental heart failure.

Inhibitors of hydroxymethylglutaryl-CoA reductase or statins have been shown to alleviate endothelial dysfunction. Their effects on constitutive nitric oxide synthase in the central nervous system may hypothetically affect the autonomic balance in sympathoexcitatory states, such as chronic heart failure (CHF). To address this issue, simvastatin (SIM) (0.3, 1.5, or 3 mg. kg-1. day-1 po) was given to rabbits with pacing-induced CHF over a 3-wk period. Normal and CHF vehicle-treated rabbits served as controls. Autonomic balance was assessed by measuring heart rate variability, including power spectral analysis (PSA). In addition, changes in resting heart rate were assessed before and after vagal and sympathetic autonomic blockade by atropine and metoprolol, respectively. The SD for all intervals was 8.9 +/- 0.7 ms in normal, 4.9 +/- 0.6 ms in CHF (P < 0.01), 3.8 +/- 0.6 ms in CHF with 0.3 mg. kg-1. day-1 SIM (P < 0.001), 5.7 +/- 0.9 in CHF with 1.5 mg. kg-1. day-1 SIM (P < 0.05), and 7.2 +/- 0.5 in CHF with 3.0 mg. kg-1. day-1 SIM. Similarly, total power was 40.5 +/- 6.3 ms2 in normal, 10.1 +/- 3.0 ms2 in CHF (P < 0.01), 6.0 +/- 1.6 ms2 in CHF with 0.3 mg. kg-1. day-1 SIM (P < 0.01), 13.2 +/- 3.9 ms2 in CHF with 1.5 mg. kg-1. day-1 SIM (P < 0.05), and 22.0 +/- 3.0 ms2 in CHF with 3.0 mg. kg-1. day-1 SIM. Both PSA data for low (0.625-0.1875 Hz) and high frequencies (0.1875-0.5625 Hz) showed recovery in CHF animals on medium and high SIM doses without changes in the low-to-high-frequency ratio. SIM beneficially affects autonomic tone in CHF as seen by the reversal of depressed HRV and total power of PSA. These data have important implications for the treatment of patients with autonomic imbalance.     

Diaphragm single-fiber weakness and loss of myosin in congestive heart failure rats

Diaphragm weakness commonly occurs in patients with congestive heart failure (CHF) and is an independent predictor of mortality. However, the pathophysiology of diaphragm weakness is poorly understood. We hypothesized that CHF induces diaphragm weakness at the single-fiber level by decreasing myosin content. In addition, we hypothesized that myofibrillar Ca(2+) sensitivity is decreased and cross-bridge kinetics are slower in CHF diaphragm fibers. Finally, we hypothesized that loss of myosin in CHF diaphragm weakness is associated with increased proteolytic activities of caspase-3 and the proteasome. In skinned diaphragm single fibers of rats with CHF, induced by left coronary artery ligation, maximum force generation was reduced by approximately 35% (P < 0.01) compared with sham-operated animals for slow, 2a, and 2x fibers. In these CHF diaphragm fibers, myosin heavy chain content per half-sarcomere was concomitantly decreased (P < 0.01). Ca(2+) sensitivity of force generation and the rate constant of tension redevelopment were significantly reduced in CHF diaphragm fibers compared with sham-operated animals for all fiber types. The cleavage activity of the proteolytic enzyme caspase-3 and the proteasome were approximately 30% (P < 0.05) and approximately 60% (P < 0.05) higher, respectively, in diaphragm homogenates from CHF rats than from sham-operated rats. The present study demonstrates diaphragm weakness at the single-fiber level in a myocardial infarct model of CHF. The reduced maximal force generation can be explained by a loss of myosin content in all fiber types and is associated with activation of caspase-3 and the proteasome. Furthermore, CHF decreases myofibrillar Ca(2+) sensitivity and slows cross-bridge cycling kinetics in diaphragm fibers.     

Low-intensity exercise training delays onset of decompensated heart failure in spontaneously hypertensive heart failure rats

Data regarding the effectiveness of chronic exercise training in improving survival in patients with congestive heart failure (CHF) are inconclusive. Therefore, we conducted a study to determine the effect of exercise training on survival in a well-defined animal model of heart failure (HF), using the lean male spontaneously hypertensive HF (SHHF) rat. In this model, animals typically present with decompensated, dilated HF between approximately 18 and 23 mo of age. SHHF rats were assigned to sedentary or exercise-trained groups at 9 and 16 mo of age. Exercise training consisted of 6 mo of low-intensity treadmill running. Exercise training delayed the onset of overt HF and improved survival (P < 0.01), independent of any effects on the hypertensive status of the rats. Training delayed the myosin heavy chain (MyHC) isoform shift from alpha- to beta-MyHC that was seen in sedentary animals that developed HF. Exercise was associated with a concurrent increase in cardiomyocyte length (approximately 6%), width, and area and prevented the increase in the length-to-width ratio seen in sedentary animals in HF. The increases in proteinuria, plasma atrial natriuretic peptide, and serum leptin levels observed in rats with HF were suppressed by low-intensity exercise training. No significant alterations in sarco(endo)plasmic reticulum Ca2+ ATPase, phospholamban, or Na+/Ca2+ exchanger protein expression were found in response to training. Our results indicate that 6 mo of low-intensity exercise training delays the onset of decompensated HF and improves survival in the male SHHF rat. Similarly, exercise intervention prevented or suppressed alterations in several key variables that normally occur with the development of overt CHF. These data support the idea that exercise may be a useful and inexpensive intervention in the treatment of HF.