Human Dilated Cardiomyopathy Research Articles

Datp Restores the Contractile Function of Cardiac Myofibril from Adult Dogs with Naturally Occurring Dilated Cardiomyopathy

Dilated cardiomyopathy (DCM) is a major type of heart failure that usually results in loss of systolic function. Naturally-occurring canine DCM is a widely-accepted experimental paradigm for studying human DCM. 2-Deoxy-adenosine triphosphate (dATP) has been demonstrated as a superior energy-substrate over ATP for myosin cross-bridge formation and increased systolic function. Here, we report the first observations on the contractile properties of isolated cardiac myofibrils from both non-failing (NF) and DCM canine samples, and evaluated the beneficial effects of dATP on contractile performance of DCM canine samples.Compared to NF samples, tension was lower for DCM myofibrils at all measured Ca2+ levels, and pCa50 was right shifted by 0.22 pCa units. NTPase assays indicated replacement of ATP with dATP significantly increased myofilament activity in both NF and DCM samples. With substitution of dATP for ATP on DCM canine myofibrils, maximal tension (64.9±2.0 vs. 50.9±1.5 mN/mm2, p<0.005) and the Ca2+ sensitivity of tension was significantly improved for DCM myofibrils, restoring them to NF levels. dATP significantly increased the kinetics of activation (kACT), but had little impact on the rate of tension redevelopment (kTR), thus restoring the ratio of kACT/kTR (0.67±0.03, vs. 0.50±0.02, p<0.005), to the NF level (0.73±0.04). This suggests thin filament activation kinetics is impaired with DCM and restored when dATP is the contractile substrate. Additionally, there were no effects on the time to 50% (0.34±0.01 vs. 0.32±0.01, p=0.2565) and 90% (2.66±0.09 vs. 2.50±0.11, p=0.2515) relaxation, suggesting that dATP increases contraction in naturally-occurring canine DCM myofibrils, without impairing relaxation. Our findings suggest increasing cardiomyocyte dATP has potential as a therapeutic strategy to recover reduced systolic ventricular performance of DCM myocardium and is worth further evaluation for the treatment of DCM.

The Benefits of Voluntaly Exersice on Cardiac Function in DCM Model Mice

Dilated cardiomyopathy (DCM) is a progressive disease characterized by ventricular dilatation and contractile dysfunction. DCM patients often die with chronic heart failure (50%) or lethal arrhythmia (30∼40%). To date, the exercise is considered to be one of therapy for HF, but the effects of exercise on DCM patients are still unclear. In this study, we examined effects of exercise on progression of HF and arrhythmogenicity in DCM using a DCM mouse model that closely mimics human inherited DCM.We used a knock-in mouse model having human DCM mutation, TNNT2 ΔK210, which decreases Ca2+ sensitivity of myofilaments. HomozygousΔK210 (DCM) mice and wild type (WT) mice at 1 month-old were housed with a running wheel (diameter = 12 cm) continuously or intermittently, and daily voluntary running activity was recorded. At 2 and 3 month-old, end-diastolic dimension and ejection fraction (EF) were measured by echocardiography. Heart, lung and lower extremity muscle were excised and their weights were measured together with body weight. Gene expressions of markers for HF and major ion channels were quantified by q-PCR analysis.The control DCM mice without running exercise showed enlarged heart, frequent SD with t1/2 of 70 days, and lowered systolic cardiac function defined by the average EF. DCM mice with running exercise showed significantly improved survival rate and less marked cardiac enlargement with dose dependent manner. Moreover, the average EF was higher in DCM mice with exercise. These beneficial effects seem to be related to the amount of exercise assessed from lower extremity muscle weights. We also found that down-regulation of some K+ channels were partially suppressed in DCM mice with exercise. These results suggest that voluntary exercise started at young age may be beneficial to DCM patients.

Modeling GATAD1-Associated Dilated Cardiomyopathy in Adult Zebrafish.

Animal models have played a critical role in validating human dilated cardiomyopathy (DCM) genes, particularly those that implicate novel mechanisms for heart failure. However, the disease phenotype may be delayed due to age-dependent penetrance. For this reason, we generated an adult zebrafish model, which is a simpler vertebrate model with higher throughput than rodents. Specifically, we studied the zebrafish homologue of GATAD1, a recently identified gene for adult-onset autosomal recessive DCM. We showed cardiac expression of gatad1 transcripts, by whole mount in situ hybridization in zebrafish embryos, and demonstrated nuclear and sarcomeric I-band subcellular localization of Gatad1 protein in cardiomyocytes, by injecting a Tol2 plasmid encoding fluorescently-tagged Gatad1. We next generated gatad1 knock-out fish lines by TALEN technology and a transgenic fish line that expresses the human DCM GATAD1-S102P mutation in cardiomyocytes. Under stress conditions, longitudinal studies uncovered heart failure (HF)-like phenotypes in stable KO mutants and a tendency toward HF phenotypes in transgenic lines. Based on these efforts of studying a gene-based inherited cardiomyopathy model, we discuss the strengths and bottlenecks of adult zebrafish as a new vertebrate model for assessing candidate cardiomyopathy genes.

Multiple Genetic Associations with Irish Wolfhound Dilated Cardiomyopathy

Cardiac disease is a leading cause of morbidity and mortality in dogs and humans, with dilated cardiomyopathy being a large contributor to this. The Irish Wolfhound (IWH) is one of the most commonly affected breeds and one of the few breeds with genetic loci associated with the disease. Mutations in more than 50 genes are associated with human dilated cardiomyopathy (DCM), yet very few are also associated with canine DCM. Furthermore, none of the identified canine loci explain many cases of the disease and previous work has indicated that genotypes at multiple loci may act together to influence disease development. In this study, loci previously associated with DCM in IWH were tested for associations in a new cohort both individually and in combination. We have identified loci significantly associated with the disease individually, but no genotypes individually or in pairs conferred a significantly greater risk of developing DCM than the population risk. However combining three loci together did result in the identification of a genotype which conferred a greater risk of disease than the overall population risk. This study suggests multiple rather than individual genetic factors, cooperating to influence DCM risk in IWH.

Clinical significance of cysteine cathepsins in human dilated cardiomyopathy

Clinical significance of cysteine cathepsins in human dilated cardiomyopathy

TBX20 loss-of-function mutation associated with familial dilated cardiomyopathy.

Dilated cardiomyopathy (DCM) is a major cause of congestive heart failure, sudden cardiac death and cardiac transplantation. Aggregating evidence highlights the genetic origin of DCM. However, DCM is a genetically heterogeneous disorder, and the genetic components underlying DCM in most cases remain unknown. The coding regions and splicing junction sites of the TBX20 gene were sequenced in 120 unrelated patients with idiopathic DCM. The available close relatives of the index patient carrying an identified mutation and 300 unrelated ethnically matched healthy individuals used as controls were genotyped for TBX20. The functional characteristics of the mutant TBX20 were assayed in contrast to its wild-type counterpart by using a dual-luciferase reporter assay system. A novel heterozygous TBX20 mutation, p.F256I, was identified in a family with DCM transmitted in an autosomal dominant fashion, which co-segregated with DCM in the family with complete penetrance. The missense mutation was absent in 600 control chromosomes and the altered amino acid was completely conserved evolutionarily among various species. Functional assays revealed that the mutant TBX20 had significantly diminished transcriptional activity. Furthermore, the mutation markedly reduced the synergistic activation of TBX20 with NKX2-5 or GATA4. This study links TBX20 loss-of-function mutation to idiopathic DCM in humans for the first time, providing novel insight into the molecular mechanism underpinning DCM.

Abstract 10260: Thrombin is a Novel Target of the Treatment of Dilated Cardiomyopathy

Introduction: A state of hypercoagulability has been observed in patients with dilated cardiomyopathy (DCM) compared to healthy subjects. In addition to being found in blood, thrombin is also expressed in the heart. Hypothesis: We hypothesize that thrombin in the heart tissue may contribute to the pathology of DCM and thrombin inhibition may be beneficial for the development of DCM. Methods: We evaluated the expression of thrombin by immunohistochemical analysis in the left ventricle of 5 patients with DCM undergoing Batista operation and that of 4 patients without heart disease serving as controls. The immunohistochemical staining was scored subjectively on a semi-quantitative scale of 0-4. We investigated the effects of the direct thrombin inhibitor, dabigatran, in the development of DCM in a mouse model carrying a deletion mutant of cardiac troponin T (DCM mouse) which causes human DCM, by using echocardiography and Kaplan-Meier method. We also estimated the apoptotic index using the terminal dUTP nick-end labeling (TUNEL) assay using the heart tissue from DCM mouse. Results: Immunohistochemical analysis showed strong thrombin expression in DCM patients compared to patients without heart disease [3.60 in DCM (n=5), 1.25 in control (n=4), p&lt;0.001]. Dabigatran significantly improved impaired left ventricular function of DCM mouse by echocardiographic examination; fractional shortening was 15.6±5.7 % in DCM mouse and was 42.7±7.3 % in DCM with dabigatran (p=0.03, n=4). Dabigatran also significantly improved the poor outcome of DCM mouse (p=0.02, n=7) (Figure1). Although there were no significant differences, dabigatran tended to reduce the apoptotic index; the percentage of TUNEL-positive nuclei was 0.20±0.04 % in DCM mouse and was 0.08±0.05 % in DCM with dabigatran (p=0.11, n=4). Conclusion: Upregulation of tissue thrombin may be involved in the pathogenesis of DCM. The direct thrombin inhibitor, dabigatran, may be a possible treatment option against DCM.

2-Deoxyadenosine triphosphate restores the contractile function of cardiac myofibril from adult dogs with naturally occurring dilated cardiomyopathy.

Dilated cardiomyopathy (DCM) is a major type of heart failure resulting from loss of systolic function. Naturally occurring canine DCM is a widely accepted experimental paradigm for studying human DCM. 2-Deoxyadenosine triphosphate (dATP) can be used by myosin and is a superior energy substrate over ATP for cross-bridge formation and increased systolic function. The objective of this study was to evaluate the beneficial effect of dATP on contractile function of cardiac myofibrils from dogs with naturally occurring DCM. We measured actomyosin NTPase activity and contraction/relaxation properties of isolated myofibrils from nonfailing (NF) and DCM canine hearts. NTPase assays indicated replacement of ATP with dATP significantly increased myofilament activity in both NF and DCM samples. dATP significantly improved maximal tension of DCM myofibrils to the NF sample level. dATP also restored Ca(2+) sensitivity of tension that was reduced in DCM samples. Similarly, dATP increased the kinetics of contractile activation (kACT), with no impact on the rate of cross-bridge tension redevelopment (kTR). Thus, the activation kinetics (kACT/kTR) that were reduced in DCM samples were restored for dATP to NF sample levels. dATP had little effect on relaxation. The rate of early slow-phase relaxation was slightly reduced with dATP, but its duration was not, nor was the fast-phase relaxation or times to 50 and 90% relaxation. Our findings suggest that myosin utilization of dATP improves cardiac myofibril contractile properties of naturally occurring DCM canine samples, restoring them to NF levels, without compromising relaxation. This suggests elevation of cardiac dATP is a promising approach for the treatment of DCM.

167 The pathogenic implications of prelamin a accumulation in cardiomyocytes: a model of premature cardiac senescence?

Introduction Mutations in the LMNA gene, which encodes the nuclear intermediate filament proteins lamins A and C, lead to a number of premature ageing syndromes, including Hutchinson Gilford Progeria syndrome (HGPS) and Emery Dreifuss muscular dystrophy (EDMD) as well as dilated cardiomyopathy (DCM). Some causal mutations disrupt lamin A processing, resulting in the accumulation of the lamin A precursor, prelamin A, however it is not clear to what extent accumulated prelamin A contributes to DCM. Here we identify DCM patients with cardiomyocyte accumulation of prelamin A and explore the impact of prelamin A accumulation in vivo . Methods We generated a novel line of targeted transgenic mice that accumulate prelamin A specifically in cardiomyocytes (PLA Tg mice), by expressing an uncleavable form of prelamin A, driven by Cre expression from the myosin light chain 2 ventricular (MLC2v) promoter. Results Immunofluorescence staining of human DCM biopsies showed the presence of nuclear prelamin A in cardiomyocytes. PLA Tg mice were born without any obvious phenotype but manifested retarded growth by 3 weeks of age and succumbed to heart failure at ˜5 weeks. At 4 weeks, MRI showed a marked dilatation of the cardiac chambers and a decline in cardiac function in vivo . Ejection fraction (EF) was substantially depressed in PLA Tg micecompared with wildtype indicating DCM and heart failure. Cardiac histology showed marked cardiomyocyte disarray and profound fibrosis. Biochemical and microscopic analyses indicated disruption of the linkers of nucleoskeleton to cytoskeleton complex and perinuclear intermediate filament network and was supported by electron micrographs showing nuclear morphology defects. Myocardial infiltration of CD45 positive cells coincided with the expression of γ-H2AX, phosphorylated ATM and increased NF-κB signalling, which suggested an inflammatory response initiated by DNA damage. This may be related to the senescence associated secretory phenotype (SASP) as senescence associated β-galactosidase was also expressed in PLA Tg myocardium. Conclusion We have demonstrated prelamin A accumulation in the nuclei of human DCM biopsies and show that overexpression of prelamin A in a transgenic mouse model leads to an early decline in cardiac function and premature myocardial senescence.

Phosphorylation state-dependent interaction between AKAP7δ/γ and phospholamban increases phospholamban phosphorylation

Changes in heart rate and contractility in response to sympathetic stimulation occur via activation of cAMP dependent protein kinase A (PKA), leading to phosphorylation of numerous substrates that alter Ca2+ cycling. Phosphorylation of these substrates is coordinated by A-kinase anchoring proteins (AKAPs), which recruit PKA to specific substrates [1]. Phosphorylation of the PKA substrate phospholamban (PLB) is a critical determinant of Ca2+ re-entry into the sarcoplasmic reticulum and is coordinated by AKAP7δ/γ [2,3]. Here, we further these findings by showing that phosphorylation of PLB requires interaction with AKAP7δ/γ and that this interaction occurs only when PLB is unphosphorylated. Additionally, we find that two mutants of PLB (R9C and Δ14), which are associated with dilated cardiomyopathy in humans, prevent association with AKAP7δ/γ and display reduced phosphorylation in vitro. This finding implicates the AKAP7δ/γ–PLB interaction in the pathology of the disease phenotype. Further exploration of the AKAP7δ/γ–PLB association demonstrated a phosphorylation state-dependence of the interaction. Computational modeling revealed that this mode of interaction allows for small amounts of AKAP and PKA (100–200nM) to regulate the phosphorylation of large quantities of PLB (50μM). Our results confirm that AKAP7γ/δ binding to PLB is important for phosphorylation of PLB, and describe a novel phosphorylation state-dependent binding mechanism that explains how phosphorylation of highly abundant PKA substrates can be regulated by AKAPs present at ~100–200 fold lower concentrations.

Prevalence and spectrum of LRRC10 mutations associated with idiopathic dilated cardiomyopathy.

Dilated cardiomyopathy (DCM) is the most common form of primary myocardial disease. It is the most common cause of chronic congestive heart failure and the most frequent reason for heart transplantation in young adults. There is increasing evidence demonstrating that genetic defects are involved in the pathogenesis of idiopathic DCM. Recent studies have shown that genetically defective LRRC10 predisposes animals to DCM. However, the association of LRRC10 with DCM in humans has not been reported. In the current study, the whole coding region and flanking splice junction sites of the LRRC10 gene were sequenced in 220 unrelated patients with idiopathic DCM. The available relatives of the index patients harboring identified mutations and 200 unrelated ethnically matched healthy individuals used as controls were also genotyped for LRRC10. The functional effect of the LRRC10 mutations was analyzed in silico. As a result, two novel heterozygous LRRC10 mutations, p.L41V and p.L163I, were identified in two families with DCM, respectively, with a mutational prevalence of ~0.91%. Genetic analyses of the pedigrees showed that in each family, the mutation co-segregated with DCM was transmitted as an autosomal dominant trait with complete penetrance. The missense mutations were absent in 400 control chromosomes and the altered amino acids were completely conserved evolutionarily across various species. Functional analysis in silico indicated that the LRRC10 mutations were causative. This study firstly reports the association of LRRC10 mutations with enhanced susceptibility to DCM in humans, which provides novel insight into the molecular mechanism underpinning DCM, and contributes to the development of novel prophylactic and therapeutic strategies for DCM.

T-tubule disease: Relationship between t-tubule organization and regional contractile performance in human dilated cardiomyopathy

Evidence from animal models suggest that t-tubule changes may play an important role in the contractile deficit associated with heart failure. However samples are usually taken at random with no regard as to regional variability present in failing hearts which leads to uncertainty in the relationship between contractile performance and possible t-tubule derangement. Regional contraction in human hearts was measured by tagged cine MRI and model fitting. At transplant, failing hearts were biopsy sampled in identified regions and immunocytochemistry was used to label t-tubules and sarcomeric z-lines. Computer image analysis was used to assess 5 different unbiased measures of t-tubule structure/organization. In regions of failing hearts that showed good contractile performance, t-tubule organization was similar to that seen in normal hearts, with worsening structure correlating with the loss of regional contractile performance. Statistical analysis showed that t-tubule direction was most highly correlated with local contractile performance, followed by the amplitude of the sarcomeric peak in the Fourier transform of the t-tubule image. Other area based measures were less well correlated. We conclude that regional contractile performance in failing human hearts is strongly correlated with the local t-tubule organization. Cluster tree analysis with a functional definition of failing contraction strength allowed a pathological definition of ‘t-tubule disease’. The regional variability in contractile performance and cellular structure is a confounding issue for analysis of samples taken from failing human hearts, although this may be overcome with regional analysis by using tagged cMRI and biopsy mapping.

A predictive model for canine dilated cardiomyopathy-a meta-analysis of Doberman Pinscher data.

Dilated cardiomyopathy is a prevalent and often fatal disease in humans and dogs. Indeed dilated cardiomyopathy is the third most common form of cardiac disease in humans, reported to affect approximately 36 individuals per 100,000 individuals. In dogs, dilated cardiomyopathy is the second most common cardiac disease and is most prevalent in the Irish Wolfhound, Doberman Pinscher and Newfoundland breeds. Dilated cardiomyopathy is characterised by ventricular chamber enlargement and systolic dysfunction which often leads to congestive heart failure. Although multiple human loci have been implicated in the pathogenesis of dilated cardiomyopathy, the identified variants are typically associated with rare monogenic forms of dilated cardiomyopathy. The potential for multigenic interactions contributing to human dilated cardiomyopathy remains poorly understood. Consistent with this, several known human dilated cardiomyopathy loci have been excluded as common causes of canine dilated cardiomyopathy, although canine dilated cardiomyopathy resembles the human disease functionally. This suggests additional genetic factors contribute to the dilated cardiomyopathy phenotype.This study represents a meta-analysis of available canine dilated cardiomyopathy genetic datasets with the goal of determining potential multigenic interactions relating the sex chromosome genotype (XX vs. XY) with known dilated cardiomyopathy associated loci on chromosome 5 and the PDK4 gene in the incidence and progression of dilated cardiomyopathy. The results show an interaction between known canine dilated cardiomyopathy loci and an unknown X-linked locus. Our study is the first to test a multigenic contribution to dilated cardiomyopathy and suggest a genetic basis for the known sex-disparity in dilated cardiomyopathy outcomes.

Bmi1 limits dilated cardiomyopathy and heart failure by inhibiting cardiac senescence.

Dilated cardiomyopathy (DCM) is the most frequent cause of heart failure and the leading indication for heart transplantation. Here we show that epigenetic regulator and central transcriptional instructor in adult stem cells, Bmi1, protects against DCM by repressing cardiac senescence. Cardiac-specific Bmi1 deletion induces the development of DCM, which progresses to lung congestion and heart failure. In contrast, Bmi1 overexpression in the heart protects from hypertrophic stimuli. Transcriptome analysis of mouse and human DCM samples indicates that p16INK4a derepression, accompanied by a senescence-associated secretory phenotype (SASP), is linked to severely impaired ventricular dimensions and contractility. Genetic reduction of p16INK4a levels reverses the pathology of Bmi1-deficient hearts. In parabiosis assays, the paracrine senescence response underlying the DCM phenotype does not transmit to healthy mice. As senescence is implicated in tissue repair and the loss of regenerative potential in aging tissues, these findings suggest a source for cardiac rejuvenation.

The functional effect of dilated cardiomyopathy mutation (R144W) in mouse cardiac troponin T is differently affected by α- and β-myosin heavy chain isoforms.

Given the differential impact of α- and β-myosin heavy chain (MHC) isoforms on how troponin T (TnT) modulates contractile dynamics, we hypothesized that the effects of dilated cardiomyopathy (DCM) mutations in TnT would be altered differently by α- and β-MHC. We characterized dynamic contractile features of normal (α-MHC) and transgenic (β-MHC) mouse cardiac muscle fibers reconstituted with a mouse TnT analog (TnTR144W) of the human DCM R141W mutation. TnTR144W did not alter maximal tension but attenuated myofilament Ca(2+) sensitivity (pCa50) to a similar extent in α- and β-MHC fibers. TnTR144W attenuated the speed of cross-bridge (XB) distortion dynamics (c) by 24% and the speed of XB recruitment dynamics (b) by 17% in α-MHC fibers; however, both b and c remained unaltered in β-MHC fibers. Likewise, TnTR144W attenuated the rates of XB detachment (g) and tension redevelopment (ktr) only in α-MHC fibers. TnTR144W also decreased the impact of strained XBs on the recruitment of new XBs (γ) by 30% only in α-MHC fibers. Because c, b, g, ktr, and γ are strongly influenced by thin filament-based cooperative mechanisms, we conclude that the TnTR144W- and β-MHC-mediated changes in the thin filament interact to produce a less severe functional phenotype, compared with that brought about by TnTR144W and α-MHC. These observations provide a basis for lower mortality rates of humans (β-MHC) harboring the TnTR141W mutant compared with transgenic mouse studies. Our findings strongly suggest that some caution is necessary when extrapolating data from transgenic mouse studies to human hearts.

Cardiac Transcriptome and Dilated Cardiomyopathy Genes in Zebrafish

Genetic studies of cardiomyopathy and heart failure have limited throughput in mammalian models. Adult zebrafish have been recently pursued as a vertebrate model with higher throughput, but genetic conservation must be tested. We conducted transcriptome analysis of zebrafish heart and searched for fish homologues of 51 known human dilated cardiomyopathy-associated genes. We also identified genes with high cardiac expression and genes with differential expression between embryonic and adult stages. Among tested genes, 30 had a single zebrafish orthologue, 14 had 2 homologues, and 5 had ≥3 homologues. By analyzing the expression data on the basis of cardiac abundance and enrichment hypotheses, we identified a single zebrafish gene for 14 of 19 multiple-homologue genes and 2 zebrafish homologues of high priority for ACTC1. Of note, our data suggested vmhc and vmhcl as functional zebrafish orthologues for human genes MYH6 and MYH7, respectively, which are established molecular markers for cardiac remodeling. Most known genes for human dilated cardiomyopathy have a corresponding zebrafish orthologue, which supports the use of zebrafish as a conserved vertebrate model. Definition of the cardiac transcriptome and fetal gene program will facilitate systems biology studies of dilated cardiomyopathy in zebrafish.

Genetics of Human and Canine Dilated Cardiomyopathy.

Cardiovascular disease is a leading cause of death in both humans and dogs. Dilated cardiomyopathy (DCM) accounts for a large number of these cases, reported to be the third most common form of cardiac disease in humans and the second most common in dogs. In human studies of DCM there are more than 50 genetic loci associated with the disease. Despite canine DCM having similar disease progression to human DCM studies into the genetic basis of canine DCM lag far behind those of human DCM. In this review the aetiology, epidemiology, and clinical characteristics of canine DCM are examined, along with highlighting possible different subtypes of canine DCM and their potential relevance to human DCM. Finally the current position of genetic research into canine and human DCM, including the genetic loci, is identified and the reasons many studies may have failed to find a genetic association with canine DCM are reviewed.

Adaptations of cytoarchitecture in human dilated cardiomyopathy.

Hypertrophic cardiomyopathy is characterised by a histological phenotype of myocyte disarray, but heart tissue samples from patients with dilated cardiomyopathy (DCM) often look comparatively similar to those from healthy individuals apart from conspicuous regions of fibrosis and necrosis. We have previously investigated subcellular alterations in the cytoarchitecture of mouse models of dilated cardiomyopathy and found that both the organisation and composition of the intercalated disc, i.e. the specialised type of cell–cell contact in the heart, is altered. There is also is a change in the composition of the M-band of the sarcomere due to an expression shift towards the more extensible embryonic heart (EH)-myomesin isoform. Analysis of human samples from the Sydney Human Heart Tissue Bank have revealed similar structural findings and also provided evidence for a dramatic change in overall cardiomyocyte size control, which has also been seen in the mouse. Together these changes in cytoarchitecture probably contribute to the decreased functional output that is seen in DCM.

Six1 is down-regulated in end-stage human dilated cardiomyopathy independently of Ezh2.

BackgroundRecently, it was shown that a knock‐out (KO) of the polycomb histone methyltransferase Ezh2 leads to cardiac hypertrophy in mice, which was driven by the homeodomain transcription factor Six1. Here, we analyzed the expression of Six1 and its regulating factor Ezh2 in cardiac tissue of patients with end‐stage dilative cardiomyopathy (DCM).MethodsTissue samples of patients with end‐stage DCM (n = 35) were compared with controls (n = 12) for the protein expression of Ezh1, Ezh2, Six1, and a marker of protein expression p70S6K.ResultsContrary to the Ezh2‐KO mouse model, we found a down‐regulation of Six1 (26%) and an up‐regulation of Ezh2 (76%) in DCM hearts, (both P < 0.05). Expression of Ezh2 and Six1 did not correlate in human tissue (DCM: r 2: 0.03, P = 0.31 and donor: r 2: 0.05, P = 0.45). Expression of Six1 weakly correlated with left ventricular end‐systolic diameter and fractional shortening. In DCM, Six1 also showed a positive correlation to the expression of the ribosomal protein p70S6K (r: 0.39, P = 0.029), which is involved in protein synthesis. This correlation was not seen in donor tissue, which showed a trend for a negative correlation (r: −0.49, P = 0.08).ConclusionOur data indicate that the Ezh2/Six1 axis might be involved in human DCM. However, Six1 expression may be regulated by factors other than Ezh2, and more research is needed to determine the precise role of Ezh2/Six1 in human DCM.

Abstract 11977: Tranilast, Transient Receptor Potential Vanilloid 2 Antagonist, Ameliorates End-Stage Heart Failure of Mice With Dilated Cardiomyopathy

Introduction: Abnormal intracellular Ca2+ handling seems to be involved in the pathogenesis of idiopathic dilated cardiomyopathy (DCM). We have found up-regulation of the expression of transient receptor potential vanilloid 2 (TRPV2), a calcium-permeable cation channel, in the sarcolemma of myocardium of animal and human DCM. Hypothesis: We hypothesized an orally active TRPV2 antagonist, tranilast, ameliorated heart failure symptoms of DCM mice. Methods: We used 4C30 mice (created by National Institute of Biomedical Innovation, Japan), which has abnormal myocardial calcium handling, as a model of DCM. Sixteen 4C30 mice of 25 weeks old with end-stage heart failure were given no drug (control) or 20 mg/kg/day of carvediol (group C) or 400 mg/kg/day of tranilast (group T) or both of them (group B) for 2 weeks. Results: Blood pressure and heart rate were similar among the 4 groups. Echocardiography demonstrated tranilast improved fractional shortening (in %). Control: 6.2±2.5; Group C: 14.2±5.6, NS; Group T: 16.6±2.3, p&lt;0.05; Group B: 17.2±3.2, p&lt;0.05. Tranilast also improved cardiac hypertrophy measured with heart-to-body weight ratio (HW/BW in mg/g). Control: 12.4±2.5; Group C: 11.5±3.4, NS; Group T: 8.6±0.9, p&lt;0.05; Group B: 5.9±1.1, p&lt;0.01. Sarcolemmal expression of TRPV2 measured with immunostaining in 4C30 mice increased twice as much as syngeneic C57BL/6J. Tranilast, not carvedilol, halved the expression of TRPV2, corresponding to reduction in [Ca2+]i of isolated cardiomyocytes. Consistent with those changes, CaMKII phosphorylation reduced in the 4C30 mice after treatment of tranilast. Conclusions: Tranilast ameliorated heart failure symptoms of 4C30 mice, possibly due to the inhibition of Ca2+ influx through TRPV2.