Animal Model Of Dilated Cardiomyopathy Research Articles

Up-regulating autophagy by targeting the mTOR-4EBP1 pathway: a possible mechanism for improving cardiac function in mice with experimental dilated cardiomyopathy

BackgroundAutophagy plays a crucial role in the pathological process of cardiovascular diseases. However, little is known about the pathological mechanism underlying autophagy regulation in dilated cardiomyopathy (DCM).MethodsWe explored whether up-regulating autophagy could improve cardiac function in mice with experimental DCM through the mTOR-4EBP1 pathway. Animal model of DCM was established in BALB/c mice by immunization with porcine cardiac myosin. Both up- or down-regulation of autophagy were studied by administration of rapamycin or 3-MA in parallel. Morphology, Western blotting, and echocardiography were applied to confirm the pathological mechanisms.ResultsAutophagy was activated and autophagosomes were significantly increased in the rapamycin group. The collagen volume fraction (CVF) was decreased in the rapamycin group compared with the DCM group (9.21 ± 0.82% vs 14.38 ± 1.24%, P < 0.01). The expression of p-mTOR and p-4EBP1 were significantly decreased in rapamycin-induced autophagy activation, while the levels were increased by down-regulating autophagy with 3-MA. In the rapamycin group, the LVEF and FS were significantly increased compared with the DCM group (54.12 ± 6.48% vs 45.29 ± 6.68%, P < 0.01; 26.89 ± 4.04% vs 22.17 ± 2.82%, P < 0.05). As the inhibitor of autophagy, 3-MA aggravated the progress of maladaptive cardiac remodeling and declined cardiac function in DCM mice.ConclusionsThe study indicated a possible mechanism for improving cardiac function in mice with experimental DCM by up-regulating autophagy via the mTOR-4EBP1 pathway, which could be a promising therapeutic strategy for DCM.

Role of cardiac TBX20 in dilated cardiomyopathy.

Dilated cardiomyopathy (DCM) is an important cause of heart failure and sudden cardiac death worldwide. Transcription factor TBX20 has been shown to play a crucial role in cardiac development and maintenance of adult mouse heart. Recent studies suggest that TBX20 may have a role in pathophysiology of DCM. In the present study, we examined TBX20 expression in idiopathic DCM patients and in an animal model of cardiomyopathy, and studied its correlation with echocardiographic indices of LV function. Endomyocardial biopsies (EMBs) from intraventricular septal from the right ventricle region were obtained from idiopathic DCM patients (IDCM, n = 30) and from patients with ventricular septal defect (VSD, n = 14) with normal LVEF who served as controls. An animal model of DCM was developed by right renal artery ligation in Wistar rats. Cardiac TBX20 mRNA levels were measured by real-time PCR in IDCM, controls, and in rats. The role of DNA promoter methylation and copy number variation (CNVs) in regulating TBX20 gene expression was also investigated. Cardiac TBX20 mRNA levels were significantly increased (8.9 fold, p < 0.001) in IDCM patients and in RAL rats as compared to the control group. Cardiac TBX20 expression showed a negative correlation with LVEF (r = -0.71, p < 0.001) and a positive correlation with left ventricular end-systolic volume (r = 0.39, p = 0.038). No significant difference in TBX20 CNVs and promoter methylation was observed between IDCM patients and control group. Our results suggest a potential role of TBX20 in pathophysiology of DCM.

Lamin A/C mutations in dilated cardiomyopathy.

Dilated cardiomyopathy (DCM) is one of the leading causes of heart failure and heart transplant. Mutations in 60 genes have been associated with DCM. Approximately 6% of all DCM cases are caused by mutations in the lamin A/C gene (LMNA). LMNA codes for type-V intermediate filaments that support the structure of the nuclear membrane and are involved in chromatin structure and gene expression. Most LMNA mutations result in striated muscle diseases while the rest affects the adipose tissue, peripheral nervous system, multiple tissues or lead to progeroid syndromes/overlapping syndromes. Patients with LMNA mutations exhibit a variety of cellular and physiological phenotypes. This paper explores the current phenotypes observed in LMNA-caused DCM, the results and implications of the cellular and animal models of DCM and the prevailing theories on the pathogenesis of laminopathies.

Biomarker Exploration and Its Clinical Use

Biomarkers are useful tools as indicators/predictors of disease severity and drug responsiveness, and thus, are expected to make drug development more efficient and to accelerate proper use of approved drugs. Many academic achievements on biomarkers have been reported, but only several biomarkers are used in drug development and clinical settings. We first show our results on the pharmacogenomic analysis of the anti-cancer drug irinotecan and of Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN). UGT1A1*6 and *28 were significantly associated with altered pharmacokinetics of an irinotecan metabolite, SN-38, and with increased frequency of severe neutropenia. HLA* 58:01 and HLA-B*15:11/HLA-A*31:01 were associated with SJS/TEN by allopurinol and carbamazepine, respectively. Our papers have been cited in the package inserts of irinotecan and allopurinol. In addition to these genomic biomarkers, metabolomic biomarkers, which can reflect the disease phenotype and drug responsiveness, have been exploring for 12 major diseases in Japan, as a part of a multi-omics team with multi-national centers. In animal models of dilated cardiomyopathy and Alzheimer's disease, we found several changes in lipid metabolite levels in the diseased tissues. Moreover, two oxidized fatty acids were correlatively changed in the brain and plasma from Alzheimer's model mice before its onset, and thus, could be candidates for predictive biomarkers. Finally, we propose/discuss several key issues for academic researches on biomarker discovery and development, especially for newly coming researchers in the field of pharmaceutical sciences. We hope that this review would help novel biomarker identification and qualification in Japan.

Fenoterol enantiomers do not possess beneficial therapeutic properties of their racemic mixture in the rat model of post myocardial infarction dilated cardiomyopathy.

A salutary effect of β(2) adrenergic receptor (AR) agonist, fenoterol has been demonstrated in a rat model of post-myocardial infarction (MI) dilated cardiomyopathy (DCM). Recent reports on single cardiomyocyte experiments suggested that out of two enantiomers, RR and SS, that constitute a racemic mixture of fenoterol, only RR-enantiomer is an active component that might be a promising new drug for treatment of chronic heart failure. The objective of this study was to compare the efficacy of the RR enantiomer of fenoterol with efficacy of racemic fenoterol, and SS, an inactive enantiomer, in whole animal experimental models of DCM. Two weeks after induction of MI by permanent ligation of the anterior descending coronary artery early cardiac remodeling and MI size were assessed via echocardiography and rats were divided into treatment groups. Treatment (placebo, racemic fenoterol, RR- or SS-enantiomers of fenoterol) continued for 6 months while progression of DCM was followed by serial echocardiography. Compared with untreated rats, rats treated with racemic fenoterol demonstrated previously described attenuation of LV remodeling, functional decline and the arrest of the MI expansion during the first 2 months of treatment. On the contrary, the treatment with either RR-, or with SS-enantiomers of fenoterol was completely ineffective. The conclusion drawn on the basis of previous experiments with single cardiomyocytes that RR-enantiomer of fenoterol represents an active component of racemic fenoterol and can be further investigated as a new drug for treatment of chronic heart failure was not confirmed in the whole animal model of DCM.

Authors' reply to "Cell therapy in dilated cardiomyopathy: back to the right scientific track?"

Bias is a bad companion for scientific endeavors. In reading our review of cell therapy trials in animal models of dilated cardiomyopathy, Marin-Neto points to the diversity of animal species, methods of inducing the cardiomyopathy, cell types, techniques of injecting the cells and methods for the evaluation of the effects as weaknesses of the pre-clinical experimental approach and concludes that “no wonder that the reported results have been so variable”. If anything, all the results have been consistent in the animal models, and in contrast to what the author of the comment says, “the ultimate goals of these studies, i.e., to achieve cardioprotective effects and/or to regenerate components of the diseased heart” have clearly shown a beneficial cardiac effect and therefore do not “remain quite elusive”. Granted that the usual policy of scientific journals not to publish negative results may in itself introduce some bias into the subject; however, a careful review of the literature points to a large body of evidence showing a positive effect of cell therapy in the various experimental models of dilated cardiomyopathy. That animal models of human disease cannot faithfully reproduce the complexity and subtleties of the pathology seen in patients has been recognized by the medical and scientific community for many decades. This only adds value to the fact that different animal models have been used, ranging from mice to dogs, in the experimental setting, and justifies the need for safety and feasibility trials in well-selected patients. We agree with Marin-Neto that “the current scenario in human investigation of cell therapy for non-ischemic dilated cardiomyopathy is full of uncertainties”. Clinical research in this area has been restricted to phase I-II trials, basically concerned with the safety and feasibility of the procedure. Therefore, it is logical and expected that the scenario should still be uncertain. The clinical research roadmap requires safety studies before efficacy can be tested. In the still small number of safety clinical trials performed in dilated cardiomyopathy patients, even a smaller number have been restricted only to patients with cardiopathy of non-ischemic origin. In all but one two-case report (1), where balloon inflation for intracoronary delivery of cells was used, the procedure has been performed safely either by intracoronary or direct intramyocardial delivery in nearly 100 patients. This number is quite small if compared to the more than 1500 patients with cardiopathy of ischemic origin to whom cell therapy has already been applied, also without adverse effects related to the procedure. At any rate, the lack of events related to the immediate cell therapy procedure does not preclude that long-term adverse effects may manifest. This again is an inherent risk of any new therapy, but since the first cardiopathy patients were treated with cell therapy a little more than a decade ago, it is reasonable to assume that safety is warranted for at least the first decade. Surprisingly, in his next to last paragraph Marin-Neto tries to analyze the results of the clinical trials performed. Since all of these trials were admittedly not designed to test for efficacy his comments are unjustified. The conclusions from all of the trials is that the procedure seems to be safe and that larger, randomized trials should be performed to test for efficacy of this new therapy. But it is also unquestion-able that if some favorable results had not been observed in these small trials, larger trials would not be pursued. So what the authors of the trials did was to report the small improvements in objective and subjective variables mea-sured during the studies and that Marin-Neto took out of context. Changes in pharmacological therapies during the course of a clinical study are undesirable but are necessary for the benefit of the patient; these changes can increase or

Hemodynamics and myocardial blood flow patterns after placement of a cardiac passive restraint device in a model of dilated cardiomyopathy

The present study examined a cardiac passive restraint device which applies epicardial pressure (HeartNet Implant; Paracor Medical, Inc, Sunnyvale, Calif) in a clinically relevant model of dilated cardiomyopathy to determine effects on hemodynamic and myocardial blood flow patterns. Dilated cardiomyopatht was established in 10 pigs (3 weeks of atrial pacing, 240 beats/min). Hemodynamic parameters and regional left ventricular blood flow were measured under baseline conditions and after acute placement of the HeartNet Implant. Measurements were repeated after adenosine infusion, allowing maximal coronary vasodilation and coronary flow reserve to be determined. Left ventricular dilation and systolic dysfunction occurred relative to baseline as measured by echocardiography. Left ventricular end-diastolic dimension increased and left ventricular fractional shortening decreased (3.8 ± 0.1 vs 6.1 ± 0.2 cm and 31.6% ± 0.5% vs 16.2% ± 2.1%, both P < .05, respectively), consistent with the dilated cardiomyopathy phenotype. The HeartNet Implant was successfully deployed without arrhythmias and a computed median mid-left ventricular epicardial pressure of 1.4 mm Hg was applied by the HeartNet Implant throughout the cardiac cycle. Acute HeartNet placement did not adversely affect steady state hemodynamics. With the HeartNet Implant in place, coronary reserve was significantly blunted. In a large animal model of dilated cardiomyopathy, the cardiac passive restraint device did not appear to adversely affect basal resting myocardial blood flow. However, after acute HeartNet Implant placement, left ventricular maximal coronary reserve was blunted. These unique results suggest that cardiac passive restraint devices that apply epicardial transmural pressure can alter myocardial blood flow patterns in a model of dilated cardiomyopathy. Whether this blunting of coronary reserve holds clinical relevance with chronic passive restraint device placement remains unestablished.

Chronic Treatment With N-acetylcysteine Improves Cardiac Function but Does Not Prevent Progression of Cardiomyopathy in Syrian Cardiomyopathic Hamsters

Oxidative stress has been postulated to contribute to the onset and development of heart failure (HF). The efficacy of antioxidant therapy in HF, however, remains controversial. This study evaluates the effect of the antioxidant N-acetylcysteine (NAC, 1 g/kg per day) on cardiovascular function in 2- and 6-month-old Bio-TO2 Syrian cardiomyopathic hamsters (SCH) after treatment for 1 month and 5 months with this drug. Endothelial function, systolic blood pressure (SBP), and echocardiographic parameters were evaluated. Age-matched F1-B golden hamsters were used as controls. One month of NAC administration significantly decreased SBP in 2-month-old SCH (n = 5, P < 0.001) without modifying echocardiographic values. Five-month treatment of cardiomyopathic animals with the antioxidant improved the acetylcholine-induced relaxation in aortic rings by 24% (E( Max) value from 45.8% ± 4% to 55.3% ± 2% n = 7, P < .05) but did not modify EC(50) values for the acetylcholine concentration-response curve. In addition, 5-month administration of NAC to SCH increased ejection fraction from 39% ± 4% to 57% ± 4% (n = 11, P < .001) and decreased left ventricular end-diastolic and end-systolic volumes (from 0.38 ± 0.04 mL/100 g body weight (BW) and 0.22 ± 0.03 mL/100 g BW, before, to 0.24 ± 0.04 mL/100 g BW and 0.12 ± 0.03 mL/100 g BW after treatment, P < .01). Cardiac output index also improved after 5 months of treatment, although it did not reach statistical significance. These results suggest that antioxidant therapy alone decreases ventricular dilatation and improves cardiovascular function in this animal model of dilated cardiomyopathy, but it does not prevent the appearance of HF.

Synthetic prostacycline agonist, ONO-1301, ameliorates left ventricular dysfunction and cardiac fibrosis in cardiomyopathic hamsters

Impairment of cardiac function in cardiomyopathy has been postulated to be related to decreased blood flow and increased collagen synthesis. Administration of growth factors was reported to attenuate left ventricular (LV) remodeling and dysfunction in animal models of dilated cardiomyopathy. We previously reported that ONO-1301, a synthetic prostacyclin agonist with thromboxane-synthase inhibitory activity, promotes production of hepatocyte growth factor and vascular endothelial growth factor from various cell types and ameliorate ischemia-induced LV dysfunction in mice and pigs. We evaluated therapeutic efficacy of ONO-1301 in the Syrian hamster (TO-2), a model of genetically determined dilated cardiomyopathy. Either vehicle or a slow releasing form of ONO-1301 (ONO-1301-PLGA, 10 mg/kg/3 weeks) was administered subcutaneously every 3 weeks to TO-2 hamsters from 24 to 32 weeks of age ( n = 12 for each group). Age-matched F1B hamsters were used as a control. Plasma concentration of HGF was elevated in ONO-1301-PLGA group ( p < 0.05). Echocardiographic study demonstrated that LV fractional shortening was significantly improved in the ONO-1301-PLGA group (25 ± 4%, p < 0.01) compared with that in the vehicle group (19 ± 2%). Cardiac fibrosis was significantly reduced by ONO-1301-PLGA ( p < 0.05) as determined by Azan-Mallory staining. Capillary density of left ventricle was markedly reduced in TO-2 hamsters. ONO-1301-PLGA significantly increased capillary density in TO-2 group ( p < 0.05). ONO-1301 improved LV dysfunction and reduced cardiac fibrosis in the hamster model of dilated cardiomyopathy. ONO-1301 might hold a therapeutic potential in the treatment of dilated cardiomyopathy.

N‐acetylcysteine (NAC) Improves but does not Prevent the Appearance of Heart Failure in Syrian Cardiomyopathic Hamsters.

Oxidative stress has been postulated to contribute to the etiology of heart failure (HF). The efficacy of antioxidant therapy in HF, however, remains controversial. This study evaluates the effect of the antioxidant N‐acetylcysteine (NAC, 1g/kg/day) on cardiovascular function in 2‐ and 6‐month‐old Bio‐TO2 Syrian cardiomyopathic hamsters (SCH) after 1‐month and 5‐month treatment with this drug. Endothelial function, systolic blood pressure (SBP), and echocardiographic parameters were evaluated. One month of NAC administration normalized the increased SBP observed in 2‐month‐old SCH (n=5, P&lt;0.05). Five‐month treatment with NAC, improved 24% (P&lt;0.05) the EMax value from the Ach‐induced relaxation (from 45.8± 4 % to 56.1 ± 2 % n= 7) while did not modify EC50 or SBP. Ejection fraction increased 39±4 % (n=11, P&lt;0.05) and cardiac output index improved but did not reach statistical significance. In addition, LVEDV and LVESV were significantly reduced by NAC (P&lt;0.05), These results suggest that antioxidant therapy alone improves cardiovascular function in this animal model of dilated cardiomyopathy. The treatment, however, does not prevent the appearance of HF. Supported by NIH‐SCORE S06‐RR 08224.

The changes of the cardiac structure and function in cTnT R141W transgenic mice

Objective To establish the transgenic mouse of cTnT R141W gene to make an animal model of dilated cardiomyopathy. Methods A transgenic plasmid was constructed by inserting the cTnT R141W gene driven by the α-MHC promoter. The expression level of the gene was determined with Northern blotting. Pathologic changes were observed by light microscopy and transmission electronic microscopy and analyzed with echocardiography. The localization of the mutant human cTnT protein was detected by immunohistochemistry. The hypertrophy markers were analyzed by RT-PCR. Results Transgenic mice carrying the cTnT R141W mutation were established. The cTnT R141W was expressed by 1.5- to 2.0-fold that of the endogenous cTnT gene and was showed to assemble in the sarcomere. The transgenic heart exhibited a thinner ventricular wall and an enlarged ventricular chamber. Interstitial fibrosis and the elongated and lysed myofrils were also observed in the transgenic heart tissue. The function on EF%, FS% and movement of the ventricular wall was significantly decreased. The immature death occurred after 4 months of age and the immature death rate was 11.1% before 8 months of age in the cTnT R141W mice. The increased NPPB, ACTA1 and decreased ATP2A2 were detected in the transgenic heart. Conclusions The expression of mutant cTnT R141W in the mouse heart caused ventricular chamber enlargement, systolic dysfunction, myocardial hypertrophy, and interstitial fibrosis, suggesting that the cTnT R141W gene is a causal factor for DCM and that the cTnT R141W transgenic mouse is a useful animal model for the study of human DCM.

Massive complement activation in desmin deficient cardiomyopathy

The complement system plays an important role in the physiopathology of acute myocardial infarction and inhibitors of complement activation have been used in clinical trials. Also several studies provide evidence that complement activation is involved in atherogenesis and autoimmune myocarditis. In a genetic animal model of dilated cardiomyopathy (desmin deficiency) we observe massive complement activation/C3 deposition and upregulation of complement system components responsible for opsonization and anaphylatoxin release, in cardiac tissue.

The Roles of Selenium and Mercury in the Pathogenesis of Viral Cardiomyopathy

Research on the pathogenesis of nonischemic dilated cardiomyopathy (DCM) has largely been focused on the role of viral pathogens and altered immunity. Trace elements have only rarely been considered; however, clinical observations that trace elements influence cardiovascular disease have been made in populations with extreme dietary deficiency or occupational exposure. Recently, animal models of DCM have been used to explore interactions among trace elements, viral pathogens, and the immune system. Discovery of interactions of trace elements with causes for DCM has heightened awareness of potential contributions of environmental variables to DCM pathogenesis. This article reviews the present knowledge regarding trace elements, in particular selenium and mercury, in the pathogenesis of viral and immune-mediated DCM. Based on recent studies, the authors propose a novel paradigm for the pathogenesis of viral DCM that incorporates trace element imbalance and its interactions with the cellular physiology of viral-induced cardiomyocyte dysfunction.

Attenuation of oxidative stress and cardiac dysfunction by bisoprolol in an animal model of dilated cardiomyopathy

Oxidative stress is an important susceptibility factor for dilated cardiomyopathy. We have investigated the effects of bisoprolol, a β 1-selective adrenoceptor blocker, on oxidative stress and the development of cardiac dysfunction in a model of dilated cardiomyopathy. Male TO-2 and control hamsters at 8 weeks of age were treated with bisoprolol (5 mg/kg per day) or vehicle for 4 weeks. Treatment with bisoprolol prevented the progression of cardiac dysfunction in TO-2 hamsters. This drug did not affect the increase in NADPH oxidase activity but prevented the reduction in activity and expression of mitochondrial manganese-dependent superoxide dismutase as well as the increases in the concentrations of interleukin-1β and tumor necrosis factor-α in the left ventricle of TO-2 hamsters. Attenuation of the development of cardiac dysfunction by bisoprolol may thus result in part from normalization of the associated increases in the levels of oxidative stress and pro-inflammatory cytokines in the left ventricle.

A metabolic approach to the treatment of dilated cardiomyopathy in BIO T0–2 cardiomyopathic Syrian hamsters

Mechanisms underlying dilated cardiomyopathy (DCM) are poorly understood and effective therapy is still unavailable. The aim of this study was to examine the heart ultrastructure and dynamic of BIO T0-2 cardiomyopathic hamsters, an animal model of DCM, and to study in these animals, the effects of a co-formulation (HS12607) of propionyl-L-carnitine, coenzyme Q(10) and omega-3 fatty acids on cardiac mechanical parameters. Sarcomere length, Frank-Starling mechanism and force-frequency relations were studied on isolated ventricular papillary muscle from age-matched BIO F1B normal Syrian hamsters, BIO T0-2 control and BIO T0-2 HS12607-treated cardiomyopathic Syrian hamsters. At the optimum length to maximum active force, electron microscopy of left ventricular papillary muscle revealed that seven out of ten muscles studied showed shorter sarcomeres (1.20 +/- 0.29 microm), and the remaining three showed longer sarcomeres (2.80 +/- 0.13 microm), compared to those of normal hamsters (2.05 +/- 0.06 microm, n = 10). Severe alterations of the Frank-Starling mechanism, force-frequency relations and derivative parameters of contractile waves were also observed in vitro in the BIO T0-2 control hamsters. Long-term (8 weeks) treatment with HS12607 prevented alterations in sarcomere length in the BIO T0-2 cardiomyopathic hamsters; the Frank-Starling mechanism and force-frequency relations were also significantly (P < 0.05) improved in these hamsters. Therefore results of the present study strongly suggest the need for clinical studies on metabolic therapeutic intervention in the effort to stop the progression of dilated cardiomyopathy.

Models of dilated cardiomyopathy in small animals and novel positive inotropic therapies.

Several randomized clinical trials of vesnarinone and milrinone in patients with heart failure left disappointing results in the 1990s. Thereafter, use of positive inotropic agents has been avoided. Exceptions are the use of digitalis glycosides to treat mild-moderate heart failure and the intravenous administration of catecholamines and phosphodiesterase inhibitors in patients with acute and/or refractory heart failure. It is not, however, exactly known whether chronic enhancement of cardiac contractility indeed has harmful effects, besides increased risk of arrhythmia and mortality. We investigated the potential chronic benefit of positive inotropic modification to treat progressive cardiomyopathy and associated heart failure using a genetic complementation strategy of muscle lim-protein and phospholamban (PLN) double mutagenesis in the mouse and found clear evidence of positive effects. Subsequent somatic modification of PLN function via gene transfer with recombinant adeno-associated virus vectors in small animal models of dilated cardiomyopathy further supported the chronic benefit of enhanced cardiac function achieved in an beta-adrenergic stimulus-independent manner. This study examines current small animal models of dilated cardiomyopathy and recent multiple attempts to use these models as novel gene-based inotropic therapies.

Serotonin is a novel survival factor of cardiomyocytes: mitochondria as a target of 5-HT2B receptor signaling.

Identification of factors regulating cardiomyocyte survival and growth is important to understand the pathogenesis of congenital heart diseases. Little is known about the molecular mechanism of cardiac functions triggered by serotonin. The link between signaling circuitry of external stimuli and the mitochondrial apoptotic machinery is of wide interest in cardiac diseases. Using cultured cardiomyocytes and 5-hydroxytryptamine (5-HT)2B-receptor knockout mice as an animal model of dilated cardiomyopathy, for the first time we show that serotonin via the Gq-coupled 5-HT2B-receptor protect cardiomyocytes against serum deprivation-induced apoptosis as manifested by DNA fragmentation, nuclear chromatin condensation, and TUNEL labeling. Serotonin prevents cytochrome c release and caspase-9 and -3 activation after serum deprivation via cross-talks between phosphatidylinositol-3 kinase/Akt and extracellular signal-regulated kinase (ERK) 1/2 signaling pathways. Serotonin binding to 5-HT2B-receptor activates ERK kinases to inhibit Bax expression induced by serum deprivation. Serotonin via phosphatidylinositol-3 kinase/Akt can activate NF-kappaB that is required for the regulation of the mitochondrial adenine nucleotide translocator (ANT-1). Parallel to these observations, ultrastructural analysis in the 5-HT2B-receptor knockout mice heart revealed pronounced mitochondrial defects in addition to altered mitochondrial enzyme activities (cytochrome oxidase and succinate dehydrogenase) and ANT-1 and Bax expressions. These findings identify 5-HT as a novel survival factor targeting mitochondria in cardiomyocytes.

Hyperubiquitination of proteins in dilated cardiomyopathy.

Protein degradation by the ubiquitin-proteasome pathway plays an important role in a variety of fundamental cellular processes, including cell cycle regulation, transcription, antigen processing and muscle remodelling. Research into disorders associated with the ubiquitin-proteasome system has been mainly in the field of neurodegenerative diseases. It is however becoming increasingly apparent that defects in the system are responsible for a number of non-neurological pathologies. Based on initial observations made as part of a proteomic analysis of an animal model of dilated cardiomyopathy (DCM) which indicated increased activity of the ubiquitin-proteasome system, we sought to determine whether this system was perturbed in hearts of human DCM patients. We studied explanted hearts from 12 DCM, 9 ischaemic (IHD) and 12 unused donor hearts. Protein expression was examined using two-dimensional polyacrylamide gel electrophoresis, Western blotting and immunohistochemistry. Expression of mRNA was examined using real-time quantitative polymerase chain reaction. Ubiquitinated proteins were affinity purified using a ubiquitin-binding column and identified using peptide mass fingerprinting. All DCM hearts showed significantly higher expression of certain key enzymes of the ubiquitin-proteasome pathway. mRNA expression of ubiquitin carboxyl-terminal hydrolase (UCH) was significantly higher (5.4-fold) in DCM hearts than in control hearts. Myocytes in sections from DCM hearts stained positively for UCH, whereas control hearts were negative. Overall protein ubiquitination was increased two-fold in DCM relative to IHD hearts and five-fold relative to donor hearts. The ubiquitination of a number of distinct proteins was greatly enhanced in DCM hearts as revealed by anti-ubiquitin Western blots. A number of these proteins were identified using affinity purification and matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry.

Gene therapy prevents disruption of dystrophin-related proteins in a model of hereditary dilated cardiomyopathy in hamsters

Background: The TO-2 hamster is an animal model of dilated cardiomyopathy (DCM). It has genetic and clinical features in common with humans who carry the gene deletion or mutation of the δ-sarcoglycan (SG) gene, a component in dystrophin-related proteins (DRP). DRP stabilise the sarcolemma during cardiac contraction. We performed in vivo gene therapy of the TO-2 hamster, whose heart is defective in all four SG proteins, to determine its potential as a model for therapy for DCM. In addition to the hereditary origin, heart failure is aggravated by treatment with catecholamines and ameliorated by the administration of some kinds of β-antagonist both in humans and in TO-2 hamsters. Methods: Gene therapy for DCM was achieved by supplementing the δ-SG gene with rAAV vector and intramurally delivering rAAV-δ-SG into the cardiac apex and left ventricle. Results: This treatment resulted in: (i) a sustained and non-pathogenic expression of both the transcript and transgene of δ-SG and all other SG proteins; (ii) improvement to both morphological and physiological deterioration; and (iii) rescued prognosis compared with untreated TO-2 hamsters, and TO-2 hamsters transfected with reporter gene alone. Another acute heart-failure model was prepared by high-dose isoproterenol treatment in Wistar rats, which resulted in: (i) translocation of dystrophin, but not δ-SG, from the cardiac sarcolemma to the myoplasm; and (ii) fragmentation of dystrophin, probably due to the activation of endogenous protease(s) or proteasome(s) that contributed to muscular dystrophy-like degeneration occurring specifically in cardiomyocytes. Conclusions: Both the TO-2 hamster and the isoproterenol-treated Wistar rat models commonly experience disruption of dystrophin or DRP. Targeting the responsible gene with the use of a potent vector may provide a novel strategy for the treatment of advanced heart failure.

A rat model of dilated cardiomyopathy to investigate partial left ventriculectomy.

This study is the first to assess a small animal model of dilated cardiomyopathy (DCM) for evaluation of partial left ventriculectomy. Eighteen Dahl salt-sensitive (DS) rats were divided into three groups. Six rats were fed an 8% high-salt diet from the age of 7 weeks (Group 1), and similarly six rats from 8 weeks (Group 2) and six from 9 weeks (Group 3). Blood pressure (BP) was measured by the tail-cuff method and left ventricular (LV) dimensions by echocardiography. In Groups 1 and 2, systolic BP rose and reached 200 mmHg by the 10th to 11th week, when all rats died within a week without signs of heart failure. However, in Group 3, systolic BP gradually rose to 196+/-15 mmHg (mean +/- SD) at the age of 14 weeks, when LV end-diastolic diameter (EDD) was 6.2+/-0.4 mm (control 5.1+/-0.7 mm) and LV fractional shortening (FS) was 77+/-3% (control 68+/-3%). At the age of 25 to 30 weeks, all rats in Group 3 showed signs of congestive heart failure, systolic BP remained high, EDD markedly increased (8.7+/-0.6 mm), and LVFS decreased (38.9+/-8.1%). From this stage, rats survived for 13.7+/-5.9 days. We employed the Group 3 model for our pilot PLV study. Eight rats had PLV with a beating heart by plicating the LV area between the papillary muscle bases. Two rats died perioperatively but the rest survived (60% survival 3 weeks after PLV). Postoperatively, the rats' LVEDD decreased and FS improved significantly. Using DS rats, we developed a DCM model for investigating PLV. The model may contribute to scientific investigation of PLV.