Heart regeneration in adult MRL mice.

The reaction of cardiac tissue to acute injury involves interacting cascades of cellular and molecular responses that encompass inflammation, hormonal signaling, extracellular matrix remodeling, and compensatory adaptation of myocytes. Myocardial regeneration is observed in amphibians, whereas scar formation characterizes cardiac ventricular wound healing in a variety of mammalian injury models. We have previously shown that the MRL mouse strain has an extraordinary capacity to heal surgical wounds, a complex trait that maps to at least seven genetic loci. Here, we extend these studies to cardiac wounds and demonstrate that a severe transmural, cryogenically induced infarction of the right ventricle heals extensively within 60 days, with the restoration of normal myocardium and function. Scarring is markedly reduced in MRL mice compared with C57BL/6 mice, consistent with both the reduced hydroxyproline levels seen after injury and an elevated cardiomyocyte mitotic index of 10-20% for the MRL compared with 1-3% for the C57BL/6. The myocardial response to injury observed in these mice resembles the regenerative process seen in amphibians.

Lung volume reduction surgery restores the normal diaphragmatic length-tension relationship in emphysematous rats.

OBJECTIVE: Improved respiratory muscle function is a major effect of a lung volume reduction surgery. We studied length adaptation in rat diaphragmatic muscle in an attempt to elucidate the mechanism by which diaphragmatic function improves after this controversial operation. METHODS: We developed a model of elastase-induced emphysema and bilateral volume reduction through median sternotomy in rats. Five months after emphysema induction, maximum exchangeable lung volume was determined in intubated and anesthetized control animals and animals with emphysema. Costal diaphragmatic length was measured in vivo, and the length at which maximal twitch force is generated was determined on muscle strips in vitro. Also 5 months after elastase administration, another cohort underwent volume reduction or sham sternotomy. Five months after the operation, these animals were similarly studied. RESULTS: Lung volume was increased in emphysematous rats versus control rats (50.9 +/- 1.7 vs 45.4 +/- 1.3 mL, P =.001). Lung volume was decreased in emphysematous animals that had undergone volume reduction versus sham sternotomy (44.7 +/- 0.60 vs 49.4 +/- 1.0 mL, P =.001). In situ diaphragm length (1.99 +/- 0.04 vs 2.24 +/- 0.07 cm, P =.001) and the length at which maximal twitch force is generated (2.25 +/- 0.06 vs 2.48 +/- 0.09 cm, P =.038) were shorter in emphysematous than control animals. After volume reduction, in situ diaphragm length (2.13 +/- 0.06 vs 1.83 +/- 0.02 cm, P <.001) and the length at which maximal twitch force is generated (2.50 +/- 0.08 vs 2.27 +/- 0.06 cm, P =.013) were longer than in animals undergoing sham sternotomy. CONCLUSIONS: In this experimental model of emphysema and lung volume reduction surgery, emphysema shortens the length at which maximal twitch force is generated and shifts the diaphragmatic length-tension curve to lower lengths; volume reduction returns the length at which maximal twitch force is generated toward normal and shifts the diaphragmatic length-tension curve back to longer lengths. This restoration toward normal physiology may enable the improvement in diaphragmatic function seen after lung volume reduction surgery. The mechanism by which these length adaptations occur merits further investigation.

Ten weeks of rapid ventricular pacing creates a long-term model of left ventricular dysfunction.

OBJECTIVE: Rapid ventricular pacing produces a reliable model of heart failure. Cessation after 4 weeks of rapid ventricular pacing results in rapid normalization of left ventricular function, but the left ventricle remains persistently dilated. We present novel data that show that prolonged rapid ventricular pacing (10 weeks) creates a model of chronic left ventricular dysfunction. METHODS: In 9 dogs undergoing 10 weeks of rapid ventricular pacing, left ventricular function and volumes were serially assessed by using 2-dimensional echocardiography and pressure-volume analysis for 12 weeks after cessation of pacing. RESULTS: Increased end-diastolic volume and decreased systolic and diastolic function were seen at the end of pacing. By 2 weeks of recovery from rapid ventricular pacing, end-diastolic volume and ejection fraction were partially recovered but did not improve further thereafter. Load-independent and load-sensitive indices of function obtained by pressure-volume analysis at 8 and 12 weeks of recovery confirmed a persistence of both systolic and diastolic dysfunction. In addition, left ventricular mass increased with pacing and remained elevated at 8 and 12 weeks of recovery. Four of these dogs studied at 6 months of recovery showed similar left ventricular abnormalities. CONCLUSION: Ten weeks of rapid ventricular pacing creates a long-term model of left ventricular dysfunction.

Cardiovascular effects of inhaled nitric oxide in a canine model of cardiomyopathy.

BACKGROUND: The inhalation of nitric oxide (NO) in patients with heart failure decreases pulmonary vascular resistance (PVR) and is associated with an increase in pulmonary artery wedge pressure (PAWP). The mechanism for this effect remains unclear. METHODS: In dogs rapid-paced for 8 weeks to induce cardiac dysfunction, we performed left ventricular pressure-volume analysis of unpaced hearts in situ to determine whether during NO inhalation (80 ppm), the mechanism for the rise in PAWP is due to: 1) primary pulmonary vasodilation; 2) a direct negative inotropic effect; or 3) impairment of ventricular relaxation. RESULTS: Inhalation of NO decreased PVR by 51%+/-3.8% (257+/-25 vs 127+/-18 dynes x sec x cm(-5) [NO 80 ppm]; p < 0.001) and increased PAWP (15.4+/-2.4 vs 18.1+/-2.6 mm Hg [NO 80 ppm]; p < 0.001). Calculated systemic vascular resistance remained unchanged. Left ventricular (LV) end-diastolic pressure rose (16.4+/-1.9 vs 19.1+/-1.8 mm Hg [NO 80 ppm]; p < 0.001), as did LV end-diastolic volume (83.5+/-4.0 vs 77.0+/-3.4 mL [NO 80 ppm]; p = 0.006). LV peak +dP/dt was unchanged by NO (1,082+/-105 vs 1,142+/-111 mm Hg/sec [NO 80 ppm]; p = NS). There was a trend toward a stroke volume increase (17.4+/-1.2 vs 18.8+/-1.3 mL; p = NS), but the relaxation time constant and end-diastolic pressure-volume relation were both unchanged. CONCLUSIONS: In this canine model of cardiomyopathy, inhaled NO decreases pulmonary vascular resistance. The associated increase in left ventricular filling pressure appears to be secondary to a primary pulmonary vasodilator effect of NO without primary effects on the contractile or relaxation properties of the left ventricle.

Stable restoration of the sarcoglycan complex in dystrophic muscle perfused with histamine and a recombinant adeno-associated viral vector.

Limb-girdle muscular dystrophies 2C-F represent a family of autosomal recessive diseases caused by defects in sarcoglycan genes. The cardiomyopathic hamster is a naturally occurring model for limb-girdle muscular dystrophy caused by a primary deficiency in delta-sarcoglycan. We show here that acute sarcolemmal disruption occurs in this animal model during forceful muscle contraction. A recombinant adeno-associated virus vector encoding human delta-sarcoglycan conferred efficient and stable genetic reconstitution in the adult cardiomyopathic hamster when injected directly into muscle. A quantitative assay demonstrated that vector-transduced muscle fibers are stably protected from sarcolemmal disruption; there was no associated inflammation or immunologic response to the vector-encoded protein. Efficient gene transduction with rescue of the sarcoglycan complex in muscle fibers of the distal hindlimb was also obtained after infusion of recombinant adeno-associated virus into the femoral artery in conjunction with histamine-induced endothelial permeabilization. This study provides a strong rationale for the development of gene therapy for limb-girdle muscular dystrophy.

The stretch-activation response may be critical to the proper functioning of the mammalian heart.

The "stretch-activation" response is essential to the generation of the oscillatory power required for the beating of insect wings. It has been conjectured but not previously shown that a stretch-activation response contributes to the performance of a beating heart. Here, we generated transgenic mice that express a human mutant myosin essential light chain derived from a family with an inherited cardiac hypertrophy. These mice faithfully replicate the cardiac disease of the patients with this mutant allele. They provide the opportunity to study the stretch-activation response before the hearts are distorted by the hypertrophic process. Studies disclose a mismatch between the physiologic heart rate and resonant frequency of the cardiac papillary muscles expressing the mutant essential light chain. This discordance reduces oscillatory power at frequencies that correspond to physiologic heart-rates and is followed by subsequent hypertrophy. It appears, therefore, that the stretch-activation response, first described in insect flight muscle, may play a role in the mammalian heart, and its further study may suggest a new way to modulate human cardiac function.

Long-term dynamic cardiomyoplasty improves chronic and acute myocardial energetics in a model of left ventricular dysfunction.

BACKGROUND: We present the first long-term evaluation of myocardial energetics after dynamic cardiomyoplasty (CMP) in a model of left ventricular (LV) dysfunction. METHODS AND RESULTS: Seventeen dogs underwent rapid ventricular pacing (RVP) to create heart failure. Eight dogs were randomly selected to undergo cardiomyoplasty. All dogs continued RVP for 6 additional weeks, whereas the CMP dogs underwent a simultaneously delivered synchronized muscle wrap conditioning protocol. After termination of RVP at 10 weeks in all dogs, myoplasty dogs continued to receive muscle wrap stimulation until the terminal study. Pressure-volume analysis to assess LV energetics was conducted at baseline and 4 weeks and 3 months after termination of RVP (6 months after baseline). At 6 months, CMP dogs displayed enhanced contractility, lower volumes, and more optimal energetics compared with control animals. Acute muscle wrap stimulation further increased effective contractility and myocardial efficiency compared with unassisted beats. CONCLUSIONS: The decrease in NYHA functional class that occurs in patients after dynamic cardiomyoplasty may be secondary to its beneficial effects on long-term myocardial function, volume, and energetics.

Modified rapid ventricular pacing: a chronic model of heart failure for evaluation of new surgical therapies.

Rapid ventricular pacing (RVP) in dogs creates a well characterized model of dilated cardiomyopathy. Standard pacing protocols use RVP at 240-260 beats/min for 2-4 weeks, and result in high mortality rates if continued longer. The authors describe a modification of RVP that results in significant heart failure by 4 weeks, but can be continued for up to 10 weeks with low mortality. Nineteen mongrels underwent RVP at 215 beats/min for 10 weeks. Serial pressure-volume analysis and echocardiography were performed in this model to assess longitudinally changes in left ventricular (LV) function and volumes. The mortality rate was 10%. Significant progressive LV dysfunction with concomitant LV enlargement was observed throughout the pacing period. Finally, norepinephrine levels were elevated at the end of pacing, consistent with an activated sympathetic system. This modified RVP protocol permits long-term pacing with a low mortality rate and results in progressive heart failure throughout the pacing period. This model would be useful in the long-term evaluation of newer surgical and medical therapies of the failing heart.

Endurance exercise alters the contractile responsiveness of rat heart to extracellular Na+ and Ca2+.

PURPOSE AND METHODS: The isovolumic contractile responsiveness of left ventricular (LV) myocardium to altered extracellular [Ca2+], [Na+], and pacing frequency was examined using perfused hearts (37 degrees C) isolated from sedentary (SED) and treadmill-trained (TR) adult female rats. RESULTS: The suppressive effect of reducing perfusate free [Ca2+] to 0.7 mM on LV developed pressure (delta LVP) was greater in the TR hearts compared with SED hearts (P < 0.05). When perfusate [Na+] was reduced to 120 mM ([Ca2+] = 0.7 mM), delta LVP augmentation was greatest in the TR hearts (P < 0.05). The negative force-frequency relationship observed at physiologic [Ca2+] and [Na+] was progressively altered toward a positive force-frequency relationship with each subsequent change in perfusate [Ca2+] and [Na+] although the effect was greatest in TR hearts (P < 0.05). CONCLUSIONS: Training elicited a small but significant (P < 0.05) prolongation in the pressure development phase of contraction. Under the physiological [Ca2+], [Na+] perfusion condition, training produced an increase in the magnitude of extrasystolic potentiation of LV pressure, whereas the time constant of mechanical restitution was unaffected. Training affected neither the Ca(2+)-dependence nor the maximal capacity of [3H] ryanodine binding to LV myocardial homogenates. The simplest interpretation of [Na+] and [Ca2+] reduction experiments is that myocardial Ca2+ efflux was augmented by exercise training.

Stabilization of chronic remodeling by asynchronous cardiomyoplasty in dilated cardiomyopathy: effects of a conditioned muscle wrap.

BACKGROUND: Dynamic cardiomyoplasty is a promising new therapy for dilated cardiomyopathy. The girdling effects of a conditioned muscle wrap alone have recently been postulated to partly explain its mechanism. We investigated this effect in a canine model of chronic dilated cardiomyopathy. METHODS AND RESULTS: Twenty dogs underwent rapid ventricular pacing (RVP) for 4 weeks to create a model of dilated cardiomyopathy. Seven dogs were then randomly selected to undergo subsequent cardiomyoplasty, and all dogs had 6 weeks of additional RVP. The cardiomyoplasty group also received 6 weeks of concurrent skeletal muscle stimulation consisting of single twitches delivered asynchronously at 2 Hz to transform the wrap without active assistance. All dogs were studied by pressure-volume analysis and echocardiography at baseline and after 4 and 10 weeks of pacing. Systolic indices, including ejection fraction (EF), end-systolic elastance (Ees), and preload-recruitable stroke work (PRSW) were all increased at 10 weeks in the wrap versus controls (EF, 34.0 versus 27.1, P=.008; Ees, 1.65 versus 1.26, P=.09; PRSW, 35.9 versus 25.5, P=.001). Ventricular volumes, diastolic relaxation, and left ventricular end-diastolic pressures stabilized in the cardiomyoplasty group but continued to deteriorate in controls. Both the end-systolic and end-diastolic pressure-volume relationships shifted farther rightward in controls but remained stable in the cardiomyoplasty group. CONCLUSIONS: In addition to potential benefits from active systolic assistance, benefits from dynamic cardiomyoplasty appear to be partially accounted for by the presence of a conditioned muscle wrap alone. This conditioned wrap stabilizes the remodeling process of heart failure, arresting progressive deterioration of systolic and diastolic function.

Dynamic cardiomyoplasty: its chronic and acute effects on the failing heart.

OBJECTIVES: Dynamic cardiomyoplasty is an alternative therapy for end-stage heart failure. We investigated the mechanisms, both acute and chronic, by which a synchronously stimulated conditioned muscle wrap affects left ventricular function in a chronic canine model of dilated cardiomyopathy. METHODS: Nineteen dogs underwent rapid ventricular pacing at a rate of 215 beats/min for 4 weeks to create a model of heart failure. Eight dogs were then randomly selected to undergo cardiomyoplasty, and all dogs received 6 additional weeks of rapid ventricular pacing. The cardiomyoplasty group also received a graded muscle conditioning protocol of synchronized burst stimulation to transform the muscle wrap. All dogs were studied with pressure-volume analysis and echocardiography at baseline and after 4 and 10 weeks of rapid ventricular pacing. Data in the cardiomyoplasty group were analyzed with the stimulator off, with it augmenting every beat (1:1), and with it augmenting only every other beat (1:2). RESULTS: Stimulator "of" data at 10 weeks of rapid pacing demonstrated chronic effects by enhanced ventricular function (end-systolic elastance = 1.80 after myoplasty vs 1.17 for controls, p = 0.005) and a stabilization of volumes and composite end-systolic and end-diastolic pressure-volume relations in the cardiomyoplasty group when compared with controls. Myoplasty stimulation increased apparent contractility (preload recruitable stroke work = 31.3 for stimulator "of" vs 40.6 for stimulator 1:2 assisted beats [p < 0.05] and vs 45.4 for stimulator 1:1 [p < 0.05]). CONCLUSIONS: Benefits from dynamic cardiomyoplasty are by at least two mechanisms: (1) the girdling effects of a conditioned muscle wrap, which halts the chronic remodeling of heart failure, and (2) active systolic assistance, which augments the apparent contractility of the failing heart.

Inhibition of myocardial crossbridge cycling by hypoxic endothelial cells: a potential mechanism for matching oxygen supply and demand?

Previous studies have shown that cardiac endothelial cells release substances that influence myocardial contraction. Since PO2 is an important stimulus that modulates endothelial function, we investigated the effects of acute moderate hypoxia and reoxygenation on the release of cardioactive factors by endothelial cells. Endothelial cells cultured from several vascular beds were superfused with normoxic (equilibrated with room air; PO2, approximately 160 mm Hg) or hypoxic (PO2, 40 to 50 mm Hg) physiological buffer solution, and the superfusates were reequilibrated to a PO2 of approximately 160 mm Hg and then tested for their effects on various myocardial assays. Endothelial cell viability and buffer ionic composition were unaltered after the superfusion procedures. The superfusates of hypoxic endothelial cells induced rapid, potent, reversible inhibition of isolated cardiac myocyte contraction without reducing cytosolic Ca2+ transients. This activity was not lost after heating (95 degrees C) and was present in low molecular weight (Mr, <500) superfusate fractions. Hypoxic endothelial superfusate reduced unloaded shortening velocity of human skinned soleus muscle fibers. It markedly depressed in vitro actin motility over cardiac myosin and reduced the rate of actin-activated cardiac myosin ATPase activity but had no effect on corresponding smooth muscle myosin assays. Reoxygenation of hypoxic endothelial cells resulted in loss of this inhibitory activity. These data indicate that cultured endothelial cells respond to acute moderate hypoxia by releasing an unidentified substance(s) that inhibits myocardial crossbridge cycling, independent of Ca2+ or other second messenger signaling pathways. Such a mechanism could have important implications for the regulation of oxygen supply-demand balance in the heart and be relevant to conditions such as myocardial hibernation.

Triiodothyronine optimizes sheep ventriculoarterial coupling for work efficiency.

BACKGROUND: Triiodothyronine (T3) administration after cardiopulmonary bypass has been shown to significantly improve cardiac performance. The present study was undertaken to elucidate the effects of T3, when administered as an intravenous bolus, on both cardiac energetics and stroke work-oxygen utilization (EW/LVVO2) efficiency. METHODS: In both unstressed and stressed hearts, energetics were evaluated at baseline and 2 hours after intervention in an in vivo sheep preparation. In the first group (n = 5) sheep received saline vehicle. In the second group (n = 9) sheep received an intravenous bolus of 1.2 micrograms/kg of T3. In the third group (n = 7) sheep received a 2-hour intravenous infusion of dobutamine at a rate of 5 micrograms/kg/min. RESULTS: In the unstressed heart, T3 improved cardiac function at no cost in oxygen consumption by decreasing afterload and hence improved EW/LVVO2 efficiency. In contrast, dobutamine improved unstressed cardiac function by increasing contractility at the cost of increased oxygen consumption and thus decreased EW/LVVO2 efficiency. Triiodothyronine optimized ventriculoarterial coupling for efficiency, but dobutamine optimized coupling for maximal work. In the stressed heart, T3 again improved EW/LVVO2 efficiency, but dobutamine had the opposite effect. CONCLUSIONS: The bolus administration of T3 improves unstressed cardiac performance through optimization of ventriculoarterial coupling for EW/LVVO2 efficiency, primarily through vasodilation. Triiodothyronine also increases efficiency in the stressed heart. This study supports the use of T3 in cardiac operations to improve cardiac performance with no cost in oxygen consumption characteristic of inotropic agents.

Acute systolic and diastolic indices of left ventricular function after cardiomyoplasty in a chronic model of heart failure.

Recent experimental studies have suggested that the initial nonstimulated stage of dynamic cardiomyoplasty acutely impairs ventricular function. Those investigations were performed on normal hearts and primarily examined diastolic alterations as a result of the passive muscle wrap. The purpose of this study was to assess the acute systolic and diastolic effects of a nonstimulated muscle wrap in chronic heart failure induced by rapid ventricular pacing in canines. Pressure-volume analysis of ventricular function based on conductance catheter volume and micromanometer pressure data was used. Each animal was studied before rapid pacing, before cardiomyoplasty, and immediately after wrap. By the end of the pacing period and before wrap, left ventricular dysfunction developed in all dogs, manifested by significant deterioration of both systolic and diastolic indices of ventricular function, as well as progressive increases in left ventricular volumes. However, no further deterioration with load insensitive indices of systolic or diastolic indicators of ventricular function was found as a result of the passive muscle wrap. These results suggest that the cardiomyoplasty procedure can be safely performed on failing hearts without prohibitive acute impairment of ventricular function.

Abnormal contractile properties of muscle fibers expressing beta-myosin heavy chain gene mutations in patients with hypertrophic cardiomyopathy.

Missense mutations in the beta-myosin heavy chain (beta-MHC) gene cause hypertrophic cardiomyopathy (HCM). As normal and mutant beta-MHCs are expressed in slow-twitch skeletal muscle of HCM patients, we compared the contractile properties of single slow-twitch muscle fibers from patients with three distinct beta-MHC gene mutations and normal controls. Fibers with the 741Gly-->Arg mutation (near the binding site of essential light chain) demonstrated decreased maximum velocity of shortening (39% of normal) and decreased isometric force generation (42% of normal). Fibers with the 403Arg-->Gln mutation (at the actin interface of myosin) showed lowered force/stiffness ratio (56% of normal) and depressed velocity of shortening (50% of normal). Both the 741Gly-->Arg and 403Arg-->Gln mutation-containing fibers displayed abnormal force-velocity relationships and reduced power output. Fibers with the 256Gly-->Glu mutation (end of ATP-binding pocket) had contractile properties that were indistinguishable from normal. Thus there is variability in the nature and extent of functional impairments in skeletal fibers containing different beta-MHC gene mutations, which may correlate with the severity and penetrance of the disease that results from each mutation. These functional alterations likely constitute the primary stimulus for the cardiac hypertrophy that is characteristic of this disease.

Does volume catheter parallel conductance vary during a cardiac cycle?

Absolute left ventricular volume measurement by the conductance (volume) catheter requires subtraction of the conductance contribution from structures extrinsic to the cavity blood pool. Previously, this parallel conductance volume (Vp) has been assumed constant throughout the cardiac cycle, and the technique described for its estimation in situ yields a single value. We present a new method for parallel conductance determination that yields multiple estimates during systole, enabling an assessment of Vp variability [Vp(t)]. For isolated blood-perfused ejecting canine left ventricles with empty (vented) right ventricles, Vp(t) displayed virtually no variation throughout systole. For in situ hearts, despite the presence of other cardiac chambers, Vp(t) also displayed little variation, with no statistically significant deviation from its mean value throughout systole. Volume signal simulations found the new technique to be less sensitive to signal noise and thus more robust than the one previously published. The isolated and in situ heart data indicate that for the left ventricle, the parallel conductance is relatively constant throughout normal ejection.