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(Circulation Research. 2003;93:1095.)
© 2003 American Heart Association, Inc.

Integrative Physiology

Reversal of Chronic Molecular and Cellular Abnormalities Due to Heart Failure by Passive Mechanical Ventricular Containment

Hani N. Sabbah, Victor G. Sharov, Ramesh C. Gupta, Sudhish Mishra, Sharad Rastogi, Albertas I. Undrovinas, Pervaiz A. Chaudhry, Anastassia Todor, Takayuki Mishima, Elaine J. Tanhehco, George Suzuki

From the Department of Medicine, Division of Cardiovascular Medicine, Henry Ford Heart and Vascular Institute, Detroit, Mich.

Correspondence to Hani N. Sabbah, PhD, Director, Cardiovascular Research, Henry Ford Hospital, 2799 W Grand Blvd, Detroit, MI 48202. E-mail HSabbah1{at}hfhs.org

	Abstract

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Passive mechanical containment of failing left ventricle (LV) with the Acorn Cardiac Support Device (CSD) was shown to prevent progressive LV dilation in dogs with heart failure (HF) and increase ejection fraction. To examine possible mechanisms for improved LV function with the CSD, we examined the effect of CSD therapy on the expression of cardiac stretch response proteins, myocyte hypertrophy, sarcoplasmic reticulum Ca²⁺-ATPase activity and uptake, and mRNA gene expression for myosin heavy chain (MHC) isoforms. HF was produced in 12 dogs by intracoronary microembolization. Six dogs were implanted with the CSD and 6 served as concurrent controls. LV tissue from 6 normal dogs was used for comparison. Compared with normal dogs, untreated HF dogs showed reduced cardiomyocyte contraction and relaxation, upregulation of stretch response proteins (p21ras, c-fos, and p38 /ß mitogen-activated protein kinase), increased myocyte hypertrophy, reduced SERCA2a activity with unchanged affinity for calcium, reduced proportion of mRNA gene expression for -MHC, and increased proportion of ß-MHC. Therapy with the CSD was associated with improved cardiomyocyte contraction and relaxation, downregulation of stretch response proteins, attenuation of cardiomyocyte hypertrophy, increased affinity of the pump for calcium, and restoration of - and ß-MHC isoforms ratio. The results suggest that preventing LV dilation and stretch with the CSD promotes downregulation of stretch response proteins, attenuates myocyte hypertrophy and improves SR calcium cycling. These data offer possible mechanisms for improvement of LV function after CSD therapy.

Key Words: heart failure • myocyte hypertrophy • sarcoplasmic reticulum • myosin heavy chain

	Introduction

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Heart failure (HF) is a progressive disorder mediated through multiple signaling pathways. Once initiated, HF is characterized by increased neurohumoral activation and ventricular dilation. Although such compensatory changes are initially beneficial, over the long-term they cause adverse structural and functional changes collectively referred to as ventricular remodeling. Ventricular dilation also causes increased mechanical stress and myocardial stretch. Upregulation of stretch response proteins, such as p21ras, ¹ c-fos,^2,3 and p38 /ß mitogen-activated protein kinase (MAPK),⁴ have been shown to induce cardiomyocyte hypertrophy.

The Acorn Cardiac Support Device (CSD) has been shown to halt progressive left ventricular (LV) dilation and improve ejection fraction.^5–7 However, the mechanism(s) underlying the improved cardiac function has not been elucidated. In the present study, we tested the hypothesis that improvement in LV systolic function in dogs with HF after long-term therapy with the CSD results, in part, from downregulation of stretch response proteins, attenuation of cardiomyocyte hypertrophy,^1–4 and improvement of sarcoplasmic reticulum (SR) calcium cycling. To further understand the mechanisms for the improvement in LV systolic function, we also explored the influence of this form of therapy on the expression of cardiac - and ß-myosin heavy chain (MHC) isoforms.^8,9

	Materials and Methods

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Animal Model
The canine model of chronic HF used in this study was previously described in detail.¹⁰ Chronic LV dysfunction is produced by multiple sequential intracoronary embolization with polystyrene Latex microspheres (70 to 102 µm in diameter), which results in loss of viable myocardium. The model manifests many of the sequelae of HF observed in humans with HF, including marked depression of LV systolic and diastolic function, reduced cardiac output, increased LV filling pressures, and enhanced activity of the sympathetic nervous system.¹⁰ Moreover, this model demonstrates progression of HF long after the cessation of coronary microembolizations. In the present study, 12 healthy mongrel dogs (Marshall Farms, North Rose, NY), weighing between 21 and 31 kg, underwent serial coronary microembolizations to produce HF. Embolizations were performed 1 to 3 weeks apart and were discontinued when LV ejection fraction was between 30% and 40%. Microembolizations were performed during cardiac catheterization under general anesthesia and sterile conditions. The anesthesia regimen consisted of a combination of intravenous injection of oxymorphone (0.22 mg/kg), diazepam (0.17 mg/kg), and sodium pentobarbital (150 to 250 mg) to effect.

Study Protocol
Dogs underwent a left and right heart catheterization at baseline, before any coronary microembolizations. At 2 weeks after the last coronary microembolization, dogs underwent another left and right heart catheterization (pretreatment) while anesthetized. The 2-week period was allowed to ensure that all infarctions produced by the last microembolization were completely healed. The CSD was surgically implanted in 6 dogs as previously described.⁵ The remaining 6 dogs served as concurrent controls. All CSD-treated dogs and controls were followed up for 3 months during which time no cardioactive drugs were used. At the end of the follow-up period, a final left and right heart catheterization was performed. After the final catheterization, and while under general anesthesia, the chest was opened and the heart removed and the tissue prepared for histological and biochemical examination. LV tissue from 6 normal dogs was prepared in an identical manner and used for comparison. The study was approved by the Henry Ford Hospital Care of Experimental Animals Committee and conformed to the "Position of the American Heart Association on Research Animal Use."

Angiographic Measurements
Single-plane left ventriculograms were obtained during left heart catheterization with the dog placed on its right side. Ventriculograms were recorded on 35-mm cine film at 30 frames per second during the injection of 20 mL of contrast material (Reno-M-60, Squibb). Correction for image magnification was made with a radiopaque calibrated grid placed at the level of the LV. LV end-systolic and end-diastolic volumes (ESV and EDV, respectively) were calculated from LV silhouettes using the area-length method,¹¹ LV EF as previously described.¹⁰

Determination of Stretch Response Proteins
Expression of stretch response proteins, specifically p21ras, c-fos, and p38 /ß MAPK, was determined by Western blotting using homogenate of cardiomyocytes isolated from the LV free wall.^12,13 In parallel, expression of calsequestrin (CSQ), a protein that is not altered in HF, was also determined and used as internal control. All stretch response proteins were normalized to CSQ. The Western blot membranes were incubated with primary (p21 rabbit polyclonal IgG; p38 /ß MAPK rabbit polyclonal IgG; c-fos rabbit polyclonal IgG; all from Santa Cruz, Inc) and then with secondary (goat-anti-rabbit HRP conjugated; Chemicon) antibody for 2 hours each. The antibody-bound antigen was identified by chemiluminescence (Renaissance Western Blot Chemiluminescence Reagent, Perkin Elmer Life Sciences Inc), followed by autoradiography. The density of bands was quantified using a densitometer.

Contraction and Relaxation of Isolated Cardiomyocytes
Cardiomyocytes were isolated from the LV free wall as previously described.¹⁴ Cardiomyocyte contraction and relaxation were recorded using an edge detection algorithm.¹⁵ Contraction was evoked by electrical field stimulation at a frequency of 1.0 Hz. Percent cardiomyocyte shortening, peak velocity of shortening, and peak velocity of relengthening were measured in 5 to 10 cardiomyocytes from each dog selected at random. For each cardiomyocyte, 20 consecutive cycles were averaged to obtain a representative value, which was then used to calculate the average measures for each dog.

Determination of Cardiomyocyte Hypertrophy
Cardiomyocyte hypertrophy was determined by assessing average cardiomyocyte cross-sectional area from frozen LV tissue sections using computer-assisted planimetry.^5,16 The length and width of isolated cardiomyocytes were also determined. Isolated cardiomyocytes were visualized using a Labophot-2 Nikon microscope with objective 20. The field was transferred to a computer using a digital video camera and projected on a digital screen. The maximum length and width of approximately 1200 rod shaped cardiomyocytes from each dog were measured using computer-assisted planimetry.

Determination of SR Ca²⁺ Uptake and Cardiac SR Ca²⁺-ATPase (SERCA2a) Activity
Oxalate-dependent Ca²⁺ uptake was determined in LV homogenate as previously described.¹⁷ Briefly, an aliquot of 50 µL of 0.25 mg/mL LV homogenate was incubated at 37°C for 1 minute in 0.4 mL of Ca²⁺ uptake buffer consisting of 50 mmol/L imidazole-HCl (pH 7.0), 100 mmol/L KCl, 6 mmol/L MgCl₂, 10 mmol/L NaN₃ (included to inhibit mitochondrial Ca²⁺ uptake), 10 mmol/L potassium oxalate, 20 µmol/L ruthenium red (included to inhibit SR Ca²⁺ release), 0.5 mmol/L EGTA, and 0.01 to 10 µmol/L free Ca²⁺ (⁴⁵CaCl₂, 10 000 dpm/nmol). The reaction was initiated by adding an aliquot of 50 µL of 50 mmol/L ATP, the assay was terminated 2 minutes later, radioactivity retained on filter paper was counted, and oxalate-dependent Ca²⁺ uptake was calculated as previously described.¹⁷ SR Ca²⁺ uptake, expressed as nmol ⁴⁵Ca²⁺ sequestered/min per mg of noncollagen protein, was determined as previously described.¹⁷ For thapsigargin-sensitive SERCA2a activity measurements, membrane vesicles were prepared from LV tissue as previously described.¹⁸ SERCA2a activity was determined in the absence and presence of thapsigargin at varying calcium concentration (0.1 to 10.0 µmol/L) as previously described¹⁸ and the activity expressed as µmol Pi released/min per mg of noncollagen protein.

Determination of Expression of SERCA2a, Phospholamban (PLB), and PLB Phosphorylation
To determine SR protein levels of SERCA2a and PLB, sodium-dodecyl sulfate (SDS) extract of LV homogenate was prepared as previously described.^17,18 To freeze the phosphorylation state of the proteins, LV tissue was homogenized in the presence of the inhibitors of protein kinases (1 mmol/L EDTA, 1 mmol/L EGTA) and protein phosphatases (2 mmol/L sodium pyrophosphate and 10 mmol/L sodium fluoride). Five micrograms or the indicated amount of the SDS-extract was separated on 4% to 20% linear polyacrylamide (BioRad), transferred electrophoretically on nitrocellulose membrane, and the resulting membrane was incubated with primary antibody as previously described.^17,18 The accuracy of the electrotransfer was confirmed by staining the membrane with 0.1% amido black. Polyclonal antibodies for phosphorylated PLB at threonine-17 (Thr17) and serine-16 (Ser16) or monoclonal antibody for PLB was diluted to 500-fold or 2500-fold, respectively. Primary-antibody binding protein was visualized by incubating the blot with a second antibody, a peroxidase-conjugated anti-mouse in case of monoclonal or anti-rabbit in case of polyclonal antibodies, and the enhanced chemiluminescence assay was used as described by the supplier (Dupont-NEN). In parallel, CSQ was also determined in the LV homogenate. The intensity of the bands was quantified using a Bio-Rad model GS-670 imaging densitometer. The density of the phosphorylated PLB at Thr17 or Ser16 was normalized to the amount of PLB present in LV tissue. Protein levels of PLB and SERCA2a were normalized to CSQ. Before quantifying protein expression levels, the protein dependency of the immunodetectable bands for all proteins was established. In this study, a linear correlation was observed between densitometric units and protein content (<30 µg) for each immunodetectable protein.

Gene Expression of Cardiac - and ß-MHC
Total RNA from LV myocardium was isolated as described previously.¹⁹ Tissue samples were homogenized in RNA Stat-60 solution (150 mg tissue/1.5 mL RNA Stat 60) followed by extraction with chloroform, precipitation with isopropanol, and finally washing the precipitated RNA with 75% (v/v) ethanol. The RNA obtained was dissolved in RNase free water. The concentration of RNA was determined by spectrophotometry. Total RNA was diluted to 0.1 mg/mL concentration and denatured at 95°C for 5 minutes followed by rapid cooling in ice bath. Approximately 10 µg of total RNA was primed with 0.5 µg of oligo (dT)15 primer. Total RNA was reversed transcribed by using a cDNA synthesis kit (Promega Inc). After incubating the samples at 42°C for 1 hour, the reaction was terminated at 95°C for 5 minutes. The mRNA levels of - and ß-MHC were analyzed by amplification of cDNA by reverse transcriptase-polymerase chain reaction¹² followed by restriction enzyme digestion and then identified by agarose gel electrophoresis and ethidium bromide staining. Fluorescent bands corresponding to - and ß-MHC were quantified in densitometric units, each normalized to total MHC (-MHC+ß-MHC) and each reported as percent of total MHC.

Data Analysis
Within group comparisons between baseline, pretreatment, and posttreatment angiographic measures were made using repeated measures analysis of variance (ANOVA) with set at 0.05. If significance was attained, pairwise comparisons between groups was determined using the Student-Newman-Kuels test with a value of P<0.05 considered significant. Comparisons of biochemical measures between normal, HF controls and CSD-treated HF dogs were based on one-way analysis of variance (ANOVA) with set at 0.05. If significance was attained, pairwise comparisons between groups were determined using the Student-Newman-Kuels test with a value of P<0.05 considered significant. All data are reported as the mean±SEM.

	Results

Top Abstract Introduction Materials and Methods Results Discussion References

 
There were no significant differences at baseline and at pretreatment in EDV, ESV, or EF between dogs that were subsequently treated with the CSD and dogs assigned as concurrent controls (Table 1). After treatment, EF significantly decreased in untreated controls but increased significantly in CSD-treated dogs. This was accompanied by a significant increase in both ESV and EDV in untreated controls and by a significant reduction in both ESV and EDV in CSD-treated dogs (Table 1).

View this table: [in this window] [in a new window]   Table 1. Angiographic Measurements Obtained at Baseline (Base) Before Any Coronary Microembolizations, Before Initiating Therapy or Follow-Up (Before), and 3 Months After Initiating Therapy or Follow-Up (After)

Cardiomyocyte Contraction and Relaxation
Results of cardiomyocyte contraction and relaxation are shown in Table 2. Percent cardiomyocyte shortening, peak velocity of shortening, and peak velocity of relengthening decreased significantly in untreated HF dogs compared with normal dogs. In contrast, in dogs treated with the CSD all three measures were significantly higher than in untreated HF dogs.

View this table: [in this window] [in a new window]   Table 2. Contraction and Relaxation of Isolated Cardiomyocytes

Stretch Response Proteins
Western blots depicting changes in p21Ras, c-fos, and p38 /ß MAPK are shown in Figure 1. The summary data for all 6 dogs in each of the three groups are shown in Table 3. All three stretch response proteins, normalized to CSQ, increased significantly in untreated HF dogs compared with normal dogs. In HF dogs treated with the CSD, all three stretch response protein levels were similar to those seen in normal dogs (Figure 1, Table 3).

View larger version (24K): [in this window] [in a new window]   Figure 1. Representative Western blot for the stretch response proteins p21ras, c-fos, and p38 /ß MAPK. Implantation of the CSD significantly reduced the expression of all 3 proteins (see Table 3). NL indicates normal; HF, heart failure; and CSD, cardiac support device-treated dogs.

View this table: [in this window] [in a new window]   Table 3. Changes in Stretch Response Protein Levels, Sarcoplasmic Reticulum Ca2+ Uptake, and SERCA2a Activity, and Expression of Other Sarcoplasmic Reticulum Proteins Depicted in Densitometric Units

Cardiomyocyte Hypertrophy
Cardiomyocyte cross-sectional area increased significantly in dogs with HF compared with normal dogs. This increase was significantly attenuated by CSD treatment (Figure 2). Cardiomyocyte length and width were significantly greater in untreated HF dogs compared with normal dogs; whereas treatment with the CSD was associated with a significantly lesser change in length and width of the cardiomyocytes compared with untreated controls (Figure 2).

View larger version (21K): [in this window] [in a new window]   Figure 2. Bar graphs depicting changes in cardiomyocyte size. Treatment group abbreviations same as in Figure 1. Implantation of the CSD significantly reduced cardiomyocyte hypertrophy. MCSA indicates myocyte cross-sectional area. *P<0.05 compared with NL; **P<0.05 compared with HF.

SERCA2a Activity and Ca²⁺ Uptake
Maximal velocity (V_max) and the affinity of SERCA2a for calcium (K_0.5) are shown in Table 3. V_max, but not K_0.5, decreased significantly in control HF dogs compared with normal dogs. Therapy with the CSD did not change V_max compared with control but was associated with a significant decrease in K_0.5 compared with HF controls, indicating a higher SERCA2a affinity for calcium after CSD therapy (Table 3). V_max for SR Ca²⁺ uptake but not affinity (Ka) decreased in control HF dogs compared with normal dogs. V_max for Ca²⁺ uptake did not change after CSD therapy, whereas Ka decreased indicating an increase in the affinity after CSD therapy (Table 3).

Expression of SERCA2a, PLB, and PLB Phosphorylation
Western blots showing expression of SERCA2a, PLB, PLB at Ser16 and Thr17, and CSQ are shown in Figure 3. All proteins, with the exception of CSQ, decreased significantly in control HF dogs compared with normal dogs. Densitometric analyses in Table 3 show that expression of SERCA2a and PLB was not changed in CSD-treated dogs compared with HF controls, whereas expression of phosphorylated PLB at Ser16 and Thr17 increased with CSD therapy compared with controls.

View larger version (30K): [in this window] [in a new window]   Figure 3. Western blot showing immunodetectable sarcoplasmic reticulum proteins in left ventricular myocardium of 3 normal dogs (NL), 3 dogs with heart failure that are not treated (HF), and 3 dogs with heart failure treated with the cardiac support device (HF+CSD). SERCA2a indicates Ca2+-ATPase; PLB, phospholamban; PLB-Ser16, phosphorylated phospholamban at serine-16; PLB-Thr17, phosphorylated phospholamban at threonine-17; and CSQ, calsequestrin.

Expression of - and ß-MHC
Changes in the proportion of cardiac - and ß-MHC between normal dogs, untreated HF dogs, and CSD-treated HF dogs are shown in Table 4. In untreated HF dogs, gene expression of LV -MHC decreased significantly compared with expression in LV of normal dogs. Three months of chronic treatment with the CSD LV expression of -MHC was similar to that seen in normal dogs (Table 4). In untreated HF dogs, expression of LV ß-MHC increased significantly compared with normal dogs, whereas treatment with the CSD was associated with LV expression of ß-MHC that was similar to that seen in normal dogs (Table 4, Figure 4).

View this table: [in this window] [in a new window]   Table 4. mRNA Expression of - and ß-Myosin Heavy Chain Depicted as Percent of Total Myosin Heavy Chain

View larger version (39K): [in this window] [in a new window]   Figure 4. Ethidium bromide-agarose gel showing mRNA encoding total myosin heavy chain (MHC); -myosin heavy chain (MHC) and glyceraldehyde 1,3 diphosphate dehydrogenase (GAPDH) in LV myocardium of 3 normal dogs (NL), 3 dogs with heart failure that are not treated (HF), and 3 dogs with heart failure treated with the cardiac support device (HF+CSD).

	Discussion

Top Abstract Introduction Materials and Methods Results Discussion References

 
Heart failure is characterized by progressive LV dysfunction and dilation. Regardless of the type of initiating injury, compensatory mechanisms are evoked to maintain adequate organ perfusion that includes neurohumoral activation, ventricular dilation, and cardiomyocyte hypertrophy. These responses are beneficial initially, but in the long-term cause maladaptive changes in myocardial structural and function recognized as ventricular remodeling. Thus, sustained neurohumoral activation and increased LV mechanical stretch and wall stress associated with ventricular dilation represent key mediators that precipitate progression of HF.

In recent years, attenuation of maladaptive ventricular remodeling has become an important goal for the treatment of HF. Numerous pharmacological interventions have been developed to block various neurohumoral factors and, in doing so, attenuate remodeling. Drug therapy with angiotensin-converting enzyme inhibitors and ß-adrenergic receptor blockers represent the current standard of care in patients with HF and have been shown to attenuate LV remodeling and, in the case of ß-blockers, reverse the maladaptive process, albeit partially.^9,20 However, the existence of multiple molecular signaling pathways that can trigger HF progression suggest that even the use of multiple pharmacological agents may not completely block all pathways responsible for the progression of LV remodeling. In particular, drugs may not be as effective at blocking the effects of mechanical signals such as wall stress and myocardial stretch. The latter can have direct consequences on biochemical and molecular effector systems that can mediate LV remodeling.^1–4,25

It has long been accepted that certain surgical approaches can be combined with optimal medical therapy to provide better survival and improved quality of life in patients with advanced HF. Functional mitral regurgitation, a common feature of the failing heart, can be eliminated or attenuated by repair or replacement of the mitral valve, a procedure that can improve forward stroke output.²¹ Experience with LV assist devices (LVADs) indicates that unloading the heart can promote reduction in LV chamber size, improvement in LV performance, and normalization of gene expression.^22,23 Cardiomyoplasty is another surgical technique, in which the primary mode of action was originally thought to involve an active assist during contraction. The procedure involved wrapping a skeletal muscle around the heart and electrically stimulating the muscle to squeeze the heart and augment cardiac function. Even though the procedure involved extensive surgery and was plagued with technical difficulties, patients showed symptomatic improvement.^24,25 However, several experimental^26,27 and clinical²⁸ studies have suggested that the improvement was derived primarily from the passive girdling of the heart and not from active contraction of the skeletal muscle. Several studies in various animal models of HF have shown that progressive LV dilation can be prevented or attenuated by wrapping synthetic materials around the cardiac ventricles to elicit containment.^5–7,29 These passive mechanical devices and surgical approaches attempted to treat HF by directly preventing progressive LV enlargement and, in doing so, limit the adverse effects of increased wall stress and myocardial stretch.

The CSD is one such device designed to prevent progressive LV dilation and attenuate myocardial stretch and chamber sphericity.^5,25 Mechanical stretch has been shown to directly and/or indirectly stimulate cardiomyocyte hypertrophy through upregulation of so-called stretch response proteins.^1–4 The resulting maladaptive hypertrophy is invariably associated with abnormal SR calcium cycling, shifts in myosin isoforms, and other changes associated with ventricular remodeling.^{1–4,8,10,22,23} Thus, reducing mechanical stress and preventing excessive myocardial stretch may downregulate stretch response proteins and block an important signaling pathway for HF progression.

Findings from our laboratory and others have demonstrated that long-term monotherapy with the CSD in animals with experimentally induced HF can prevent progressive LV dilation and improve LV ejection fraction.^5–8,25 Although one would expect that a passive mechanical device such as the CSD can prevent progressive LV dilation, the mechanism by which the CSD leads to improved LV systolic function is not as clear. The present study addressed this issue by exploring the potential biochemical and molecular alterations that occurred as a consequence of CSD therapy.

In the present study, improvement of global LV function with CSD therapy was associated with lesser extent of intrinsic contractile dysfunction of cardiomyocytes compared with no treatment at all. Therapy with the CSD was also associated with lower tissue levels of stretch response proteins specifically p21ras, c-fos, and p38 /ß MAPK compared with no treatment at all. Expression of these proteins has been shown to increase in HF.^1–4 These proteins are known to be direct stimuli for cardiomyocyte hypertrophy.^1–4 Maladaptive cardiomyocyte hypertrophy plays a key role in the progression of HF.^30,31 In this study, long-term CSD therapy resulted in attenuation of cardiomyocyte hypertrophy as evidenced by decreased cardiomyocyte cross-sectional area, length, and width compared with control.

Findings of this study also showed that CSD therapy was associated with increased affinity of SERCA2a for calcium. This increase in affinity may have been due to increased phosphorylation of PLB. Increased affinity of SERCA2a for calcium can lead to improved calcium cycling within the SR particularly at low cytosolic calcium concentrations. Given that abnormalities in Ca²⁺ handling may, in part, underlie the decrease in contractile function in HF, we propose that increased affinity of the pump for calcium as seen with CSD therapy may have contributed to the observed improvement of LV function.

Marked differences in the phosphorylation of PLB were observed in the present study and warrant discussion. Phosphorylation of PLB was decreased in HF controls compared with normal dogs. It would be expected that phosphorylation of PLB would be greater in HF dogs due to the increase in plasma norepinephrine associated with the HF state. However, this increase in circulating plasma norepinephrine is accompanied by downregulation of ß₁ adrenoceptors in the heart and uncoupling between the receptors and their G proteins. In addition, phosphorylation of PLB was increased in the CSD group compared with the HF group despite the improvement of LV function, which is normally associated with decreased plasma norepinephrine. One possible explanation is that in addition to augmented plasma norepinephrine, an increase in phosphatase activity has also been documented in HF.^32–36 A decrease in PLB phosphorylation has been previously noted in our canine model of HF.^32,34 It is possible, albeit unproven, that the balance of phosphorylation/phosphatase activation may have favored dephosphorylation in the HF dogs, while reverting to phosphorylation in the CSD-treated animals.

Cardiomyocytes express both - and ß-MHC isoforms. In the rat heart, these two isoforms differ on the basis of ATPase activity, with -MHC being more active than ß-MHC.^37,38 Compared with cardiac ß-MHC, -MHC is associated with faster velocity of shortening.^37,38 Studies in LV tissue obtained form explanted failed human hearts showed loss of -MHC expression with increased expression of ß-MHC, a condition that can argue in favor of diminished contractile function. Other studies have shown that this maladaptation in the proportion of cardiac -MHC and ß-MHC isoforms can be reversed in animal models of HF after drug or surgical therapy.^39–41 In the present study, the proportion of cardiac -MHC was significantly reduced in HF dogs that were untreated, and the proportion ß-MHC was increased. Long-term treatment with the CSD was associated with expression of both MHC isoforms that was close to normal levels, a condition that may have also contributed to the improvement of LV function seen with CSD therapy.

In conclusion, results of this study suggest that the observed improvement in LV function after long-term therapy with the CSD may be due, in part, to the effects of the CSD on limiting LV wall stress and myocardial stretch. These changes were associated with attenuation of muscle cell hypertrophy and improvement of SR calcium cycling. The improvement of LV function with CSD therapy may have also been due, in part, on its effects on the expression of cardiac MHC isoforms.

	Acknowledgments

 
Acknowledgments

This study was supported, in part, by a grant from Acorn Cardiovascular, Inc, and by a grant from the National Heart, Lung, and Blood Institute (HL 49090-08).

	Footnotes

 
H.N.S. is a consultant and grant awardee for Acorn Cardiovascular, Inc.

Original received April 4, 2003; resubmission received August 5, 2003; revised resubmission received October 7, 2003; accepted October 8, 2003.

	References

Top Abstract Introduction Materials and Methods Results Discussion References

 
1. Sadoshima J, Izumo S. Mechanical stretch rapidly activates multiple signal transduction pathways in cardiac myocytes: potential involvement of an autocrine/paracrine mechanism. EMBO J. 1993; 12: 1681–1692.[Medline] [Order article via Infotrieve]

2. Sadoshima J, Izumo S. Signal transduction pathways of angiotensin II induced c-fos gene expression in cardiac myocytes in vitro: roles of phospholipid-derived second messengers. Circ Res. 1993; 73: 424–438.[Abstract/Free Full Text]

3. Sadoshima J, Jahn L, Takahashi T, Kulik TJ, Izumo S. Molecular characterization of the stretch-induced adaptation of cultured cardiac cells. An in vitro model of load-induced cardiac hypertrophy. J Biol Chem. 1992; 267: 10551–10560.[Abstract/Free Full Text]

4. Sadoshima J, Qiu Z, Morgan JP, Izumo S. Angiotensin II and other hypertrophic stimuli mediated by G protein-coupled receptors activate tyrosine kinase, mitogen-activated protein kinase, and 90-kD S6 kinase in cardiac myocytes: the critical role of Ca²⁺-dependent signaling. Circ Res. 1995; 76: 1–15.[Abstract/Free Full Text]

5. Chaudhry PA, Mishima T, Sharov VG, Hawkins J, Alferness C, Paone G, Sabbah HN. Passive epicardial containment prevents ventricular remodeling in heart failure. Ann Thorac Surg. 2000; 70: 1275–1280.[Abstract/Free Full Text]

6. Saaverda EF, Tunin RS, Paolocci N, Mishima T, Suzuki G, Emala CW, Chaudry PA, Anagnostopoulos P, Gupta RC, Sabbah HN, Kass DA. Reverse remodeling and enhanced adrenergic reserve from passive external support in experimental dilated heart failure. J Am Coll Cardiol. 2002; 39: 2069–2076.[Abstract/Free Full Text]

7. Power JM, Raman J, Dornom A, Farish SJ, Burrell LM, Tonkin AM, Buxton B, Alferness CA. Passive ventricular constraint amends the course of heart failure: a study in an ovine model of dilated cardiomyopathy. Cardiovasc Res. 1999; 44: 549–555.[Abstract/Free Full Text]

8. Nakao K, Minobe W, Roden R, Bristow MR, Leinwand LA. Myosin heavy chain gene expression in human heart failure. J Clin Invest. 1997; 100: 2362–2370.[Medline] [Order article via Infotrieve]

9. Lowes BD, Gilbert EM, Abraham WT, Minobe WA, Larrabee P, Ferguson D, Wolfel EE, Lindenfeld J, Tsvetkova T, Roberston AD, Quaife RA, Bristow MR. Myocardial gene expression in dilated cardiomyopathy treated with beta-blocking agents. N Engl J Med. 2002; 346: 1357–1365.[Abstract/Free Full Text]

10. Sabbah HN, Stein PD, Kono T, Gheorghiade M, Levine TB, Jafri S, Hawkins ET, Goldstein S. A canine model of chronic heart failure produced by multiple sequential intracoronary microembolizations. Am J Physiol. 1991; 260: H1379–H1384.[Medline] [Order article via Infotrieve]

11. Dodge HT, Sandler H, Baxley WA, Hawley RR. Usefulness and limitations of radiographic methods for determining left ventricular volume. Am J Cardiol. 1966; 18: 10–24.[CrossRef][Medline] [Order article via Infotrieve]

12. Symington J, Green M, Brackman K. Immunoautoradiographic detection of proteins after electrophoretic transfer from gels to diazo-paper: analysis of adenovirus encoded proteins. Proc Natl Acad Sci U S A. 1981; 78: 177–181.[Abstract/Free Full Text]

13. Todor A, Sharov VG, Tanhehco EJ, Silverman N, Bernabei A, Sabbah HN. Hypoxia-induced cleavage of caspase-3 and DFF45/ICAD in human failed cardiomyocytes. Am J Physiol. 2002; 283: H990–H995.

14. Maltsev VA, Sabbah HN, Higgins RSD, Silverman N, Lesch M, Undrovinas AI. Novel, ultraslow inactivating sodium current in human ventricular cardiomyocytes. Circulation. 1998; 98: 2545–2552.[Abstract/Free Full Text]

15. Maltsev VA, Sabbah HN, Tanimura M, Lesch M, Goldstein S, Undrovinas AI. Relationship between action potential, contraction-relaxation pattern, and intracellular Ca²⁺ transient in cardiomyocytes of dogs with chronic heart failure. Cell Mol Life Sci. 1998; 54: 597–605.[CrossRef][Medline] [Order article via Infotrieve]

16. Tanimura M, Sharoc VG, Shimoyama H, Mishima T, Levine TB, Goldstein S, Sabbah HN. Effects of AT₁-receptor blockade on progression of left ventricular dysfunction in dogs with heart failure. Am J Physiol. 1999; 276: H1385–H1392.[Medline] [Order article via Infotrieve]

17. Gupta RC, Mishra S, Mishima T, Goldstein S, Sabbah HN. Reduced sarcoplasmic reticulum Ca²⁺-uptake and expression of phospholamban in left ventricular myocardium of dogs with heart failure. J Moll Cell Cardiol. 1999; 31: 1381–1389.[CrossRef][Medline] [Order article via Infotrieve]

18. Gupta RC, Shimoyoma H, Tanimura M, Nair R, Lesch M, Sabbah HN. Reduced SR Ca²⁺-ATPase activity and expression in ventricular myocardium of dogs with heart failure. Am J Physiol. 1997; 273: H12–H18.[Medline] [Order article via Infotrieve]

19. Feldman AM, Ray PE, Silan CM, Mercer JA, Minobe W, Bristow MR. Selective gene expression in failing human heart: quantification of steady-state levels of messenger RNA in endomyocardial biopsies using the polymerase chain reaction. Circulation. 1991; 83: 1866–1872.[Abstract/Free Full Text]

20. Sabbah HN, Shimoyama H, Kono T, Gupta RC, Sharov VG, Scicli G, Levine B, Goldstein S. Effects of long-term monotherapy with enalapril, metoprolol, and digoxin on the progression of left ventricular dysfunction and dilation in dogs with reduced ejection fraction. Circulation. 1994; 89: 2852–2859.[Abstract/Free Full Text]

21. Bolling SF. Mitral reconstruction in cardiomyopathy. J Heart Valve Dis. 2002; 11 (suppl 1): S26–S31.[Medline] [Order article via Infotrieve]

22. Madigan JD, Barbone A, Choudhri AF, Morales DL, Cai B, Oz MC, Burkhoff D. Time course of reverse remodeling of the left ventricle during support with a left ventricular assist device. J Thorac Cardiovasc Surg. 2001; 121: 902–908.[Abstract/Free Full Text]

23. Burkhoff D, Holmes JW, Madigan J, Barbone A, Oz MC. Left ventricular assist device-induced reverse ventricular remodeling. Prog Cardiovac Dis. 2000; 43: 19–26.[CrossRef]

24. Capouya ER, Gerber RS, Drinkwater DC, Pearl JM, Sack JB, Aharon AS, Barthel SW, Kaczer EM, Chang PA, Laks H. Girdling effect of nonstimulated cardiomyoplasty on left ventricular function. Ann Thorac Surg. 1993; 56: 867–871.[Abstract]

25. Kass DA, Baughman KL, Pak PH, Cho PW, Levin HR, Gardner TJ, Halperin HR, Tsitlik JE, Acker MA. Reverse remodeling from cardiomyoplasty in human heart failure: external constraint versus active assist. Circulation. 1995; 91: 2314–2318.[Abstract/Free Full Text]

26. Vaynblat M, Chiavarelli M, Shah HR, Ramdev G, Aron M, Zisbrod Z, Cunningham JN. Cardiac binding in experimental heart failure. Ann Thorac Surg. 1997; 64: 81–85.[Abstract/Free Full Text]

27. Patel HJ, Polidori DJ, Pilla JJ, Plappert T, Kass D, St John Sutton M, Lankford EB, Acker MA. Stabilization of chronic remodeling by asynchronous cardiomyoplasty in dilated cardiomyopathy: effects of a conditioned muscle wrap. Circulation. 1997; 96: 3665–3671.[Abstract/Free Full Text]

28. Hagege AA, Desnos M, Fernandez F, Besse B, Mirochnik N, Castaldo M, Chachques JC, Carpentier A, Guerot C. Clinical study of the effects of latissimus dorsi muscle flap stimulation after cardiomyoplasty. Circulation. 1995; 92 (suppl): II-210–II-215.[Medline] [Order article via Infotrieve]

29. McCarthy PM, Takagaki M, Ochiai Y, Young JB, Tabata T, Shiota T, Qin JX, Thomas JD, Mortier TJ, Schroeder RF, Schweich J, Fukamachi K. Device-based change in left ventricular shape: a new concept for the treatment of dilated cardiomyopathy. J Thorac Cardiovasc Surg. 2001; 122: 482–490.[Abstract/Free Full Text]

30. Molkentin JD, Dorn II GW 2nd. Cytoplasmic signaling pathways that regulate cardiac hypertrophy. Annu Rev Physiol. 2001; 63: 391–426.[CrossRef][Medline] [Order article via Infotrieve]

31. Francis GS. Changing the remodeling process in heart failure: basic mechanisms and laboratory results. Curr Opin Cardiol. 1998; 13: 156–161.[Medline] [Order article via Infotrieve]

32. Gupta RC, Mishra S, Rastogi S, Imai M, Habib O, Sabbah HN. Cardiac SR-coupled type 1 protein phosphatase activity and expression are increased and inhibitor protein 1 protein expression is decreased in the failing heart. Am J Physiol Heart Circ Physiol. In press.

33. Huang B, Wang HB, Qin D, Boutjdir M, El-Sherif N. Diminished basal phosphorylation level of phospholamban in the postinfarction remodeled rat ventricle: role of ß adrenergic pathway, G_i protein, phosphodiesterase, and phosphatases. Circ Res. 1999; 85: 848–855.[Abstract/Free Full Text]

34. Mishra S, Gupta RC, Towari N, Sharov VG, Sabbah HN. Molecular mechanisms of reduced sarcoplasmic reticulum Ca²⁺ uptake in human failing left ventricular myocardium. J Heart Lung Trans. 2002; 21: 366–373.[CrossRef][Medline] [Order article via Infotrieve]

35. Sande JB, Sjaastad I, Hoen IB, Bokenes J, Tonnessen T, Holt E, Lunde PK, Christensen G. Reduced level of serine (16) phosphorylated phospholamban in the failing rat myocardium: a major contributor to reduced SERCA2 activity. Cardiovasc Res. 2002; 53: 382–391.[Abstract/Free Full Text]

36. Mishra S, Sabbah HN, Jain JC, Gupta RC. Reduced Ca²⁺-calmodulin-dependent protein kinase activity and expression in LV myocardium of dogs with heart failure. Am J Physiol Heart Circ Physiol. 2003; 284: H876–H883.[Abstract/Free Full Text]

37. Pope B, Hoh JF, Weeds A. The ATPase activities of rat cardiac myosin isoenzymes. FEBS Lett. 1980; 118: 205–208.[CrossRef][Medline] [Order article via Infotrieve]

38. Miyata S, Minobe W, Bristow MR, Leinwand LA. Myosin heavy chain isoform expression in the failing and nonfailing human heart. Circ Res. 2000; 86: 386–390.[Abstract/Free Full Text]

39. Bauersachs J, Galuppo P, Fraccarollo D, Christ M, Ertl G. Improvement of left ventricular remodeling and function by hydroxymethylglutaryl coenzyme a reductase inhibition with cerivastatin in rats with heart failure after myocardial infarction. Circulation. 2001; 104: 982–985.[Abstract/Free Full Text]

40. Liu X, Sentex E, Golfman L, Takeda S, Osada M, Dhalla NS. Modification of cardiac subcellular remodeling due to pressure overload by captopril and losartan. Clin Exp Hypertens. 1999; 21: 145–156.[CrossRef][Medline] [Order article via Infotrieve]

41. Pauletto P, Vescovo G, Scannapieco G, Angelini A, Pessina AC, Dalla Libera L, Carraro U, Dal Palu C. Changes in rat ventricular isomyosins with regression of cardiac hypertrophy. Hypertension. 1986; 8: 1143–1148.[Abstract/Free Full Text]

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