Murine Model Of Cardiomyopathy Research Articles

Simultaneous targeting of oxidative stress and fibrosis reverses cardiomyopathy-induced ventricular remodelling and dysfunction.

Oxidative stress and fibrosis are hallmarks of cardiomyopathy-induced heart failure yet are not effectively targeted by current frontline therapies. Here, the therapeutic effects of the anti-oxidant, N-acetylcysteine (NAC), were compared and combined with an acute heart failure drug with established anti-fibrotic effects, serelaxin (RLX), in a murine model of cardiomyopathy. Adult male 129sv mice were subjected to repeated isoprenaline (25 mg·kg-1 )-induced cardiac injury for five consecutive days and then left to undergo fibrotic healing until Day 14. Subgroups of isoprenaline-injured mice were treated with RLX (0.5 mg·kg-1 ·day-1 ), NAC (25 mg·kg-1 ·day-1 ) or both combined, given subcutaneously via osmotic minipumps from Day 7 to 14. Control mice received saline instead of isoprenaline. Isoprenaline-injured mice showed increased left ventricular (LV) inflammation (~5-fold), oxidative stress (~1-2.5-fold), cardiomyocyte hypertrophy (~25%), cardiac remodelling, fibrosis (~2-2.5-fold) and dysfunction by Day 14 after injury. NAC alone blocked the cardiomyopathy-induced increase in LV superoxide levels, to a greater extent than RLX. Additionally, either treatment alone only partly reduced several measures of LV inflammation, remodelling and fibrosis. In comparison, the combination of RLX and NAC prevented the cardiomyopathy-induced LV macrophage infiltration, remodelling, fibrosis and cardiomyocyte size, to a greater extent than either treatment alone after 7 days. The combination therapy also restored the isoprenaline-induced reduction in LV function, without affecting systolic BP. These findings demonstrated that the simultaneous targeting of oxidative stress and fibrosis is key to treating the pathophysiology and dysfunction induced by cardiomyopathy.

Efficacy of epicardial implantation of acellular chitosan hydrogels in ischemic and nonischemic heart failure: impact of the acetylation degree of chitosan

This work explores the epicardial implantation of acellular chitosan hydrogels in two murine models of cardiomyopathy, focusing on their potential to restore the functional capacity of the heart. Different chitosan hydrogels were generated using polymers of four degrees of acetylation, ranging from 2.5% to 38%, because the degree of acetylation affects their degradation and biological activity. The hydrogels were adjusted to a 3% final polymer concentration. After complete macromolecular characterization of the chitosans and study of the mechanical properties of the resulting hydrogels, they were sutured onto the surface of the myocardium, first in rat after four-weeks of coronary ligation (n=58) then in mice with cardiomyopathy induced by a cardiac-specific invalidation of serum response factor (n=20). The implantation of the hydrogels was associated with a reversion of cardiac function loss with maximal effects for the acetylation degree of 24%. The extent of fibrosis, the cardiomyocyte length-to-width ratio, as well as the genes involved in fibrosis and stress were repressed after implantation. Our study demonstrated the beneficial effects of chitosan hydrogels, particularly with polymers of high degrees of acetylation, on cardiac remodeling in two cardiomyopathy models. Our findings indicate they have great potential as a reliable therapeutic approach to heart failure.

Abstract P2012: Combining the Anti-Fibrotic Actions of Serelaxin With the Anti-Oxidant, N-Acetylcysteine, Abrogates Cardiac Fibrosis in an Isoprenaline-Induced Murine Model of Cardiomyopathy

Oxidative stress is a significant contributor to cardiac fibrosis, particularly in the setting of hypertension. While current frontline medications (such as ACE inhibitors) and anti-oxidants (such as N-Acetylcysteine; NAC) can attenuate the impact of elevated blood pressure (BP) on cardiac fibrosis, it is unclear to what extent established fibrosis can be reduced in the absence of BP regulation. The drug-version of the ovarian and cardiovascular hormone, relaxin, serelaxin (RLX) mediates rapid-occurring anti-fibrotic effects by inhibiting the effects of TGF-β1 on myofibroblast-induced collagen production, independently of BP regulation. In this study, we compared and combined the effects of RLX to the anti-oxidant, NAC, in a normotensive model of heart disease. 14-16 week old adult 129sv mice were subjected to isoprenaline (ISO)-induced cardiomyopathy, which along with saline-treated controls demonstrated equivalent systolic BP (SBP) measurements (122±3 vs 124±4 mmHg, respectively; as determined by tail cuff plethysmography). Sub-groups of ISO-injured mice were treated from days 7-14 post-injury with RLX (0.5mg/kg/day), NAC (25mg/kg/day; s.c) or both combined via osmotic mini-pumps. At day 14 post-injury, the left ventricle (LV) of ISO-injured mice presented with increased macrophage infiltration, phosphorylated IkB levels (a marker of NF-kB activity), oxidative stress, NOX2, NOX4 and TGF-β1 expression, myofibroblast differentiation and interstitial fibrosis (all p<0.05 vs saline-treated controls). RLX decreased cardiac fibrosis (by ~40%; p<0.05 vs ISO alone) independently of effects on macrophage infiltration, NOX2/NOX4 expression or SBP. NAC alone reduced interstitial fibrosis (by ~58%; p<0.01 vs ISO alone; no different to ISO+RLX treatment), while when added in combination, RLX and NAC completely abrogated cardiac fibrosis, returning levels to those measured in saline-treated controls. These finding suggest that NAC can reduce fibrosis without necessarily impacting on the contribution of BP to disease progression, and that the combined effects of RLX and an anti-oxidant may offer superior therapeutic efficacy in the treatment of cardiac fibrosis.

68Ga]-Albumin-PET in the Monitoring of Left Ventricular Function in Murine Models of Ischemic and Dilated Cardiomyopathy: Comparison with Cardiac MRI

The purpose of this study is to evaluate left ventricular functional parameters in healthy mice and in different murine models of cardiomyopathy with the novel blood pool (BP) positron emission tomography (PET) tracer [68Ga]-albumin. ECG-gated microPET examinations were obtained in healthy mice, and mice with dilative (DCM) and ischemic cardiomyopathy (ICM) using the novel BP tracer [68Ga]-albumin (AlbBP), as well as [18F]-FDG microPET. Cine-magnetic resonance imaging (MRI) examination performed on a clinical 1.5-T MRI provided the reference standard measurements. When considering the combined group of healthy controls, DCM and ICM AlbBP-PET significantly overestimated the magnitudes of EDV (AlbBP, 181±86 μl; cine-MRI, 125±80 μl; P<0.001) and ESV (AlbBP, 136±92 μl; cine-MRI, 96±77 μl; P<0.001), whereas the EF (AlbBP, 31±16%; cine-MRI, 33±21%; P=0.910) matched closely to cine-MRI results, as did findings with [18F]-FDG. High correlations were found between the measured cardiac parameters (EDV: R=0.978, ESV: R=0.989, and LVEF: R=0.992). Measuring left ventricular function in mice with [68Ga]-albumin BP PET is feasible and showed a high correlation compared to cine-MRI, which was used as a reference standard.

N-acetylcysteine reverses cardiac myocyte dysfunction in HIV-Tat proteinopathy

HIV cardiomyopathy remains highly prevalent among the estimated 33 million HIV-infected individuals worldwide. This is particularly true in developing countries. Potential mechanisms responsible for myocardial dysfunction following HIV infection include direct effects of HIV proteins. We have previously reported that cardiac myocyte-specific expression of HIV-Tat (Tat) results in a murine cardiomyopathy model. We now report that Tat exhibits decreased myocardial ATP [wild type (WT) vs. Tat transgenic (TG), P < 0.01] and myocyte GSH levels (WT vs. TG, P < 0.01), decreased GSH/GSSG ratio (WT vs. TG, P < 0.01), increased H(2)O(2) levels (WT vs. TG, P < 0.05), and increased catalase (TG vs. WT, P < 0.05) and GPX1 (glutathione peroxidase 1) activities (WT vs. TG, P < 0.05), blunted cardiac myocyte positive inotropy (% peak shortening, WT vs. TG, P < 0.01; +dl/dt, WT vs. TG, P < 0.01) and negative inotropy (-dl/dt, WT vs. TG, P < 0.01), and blunted inotropic responses to Ca(2+) (P < 0.01, for each) and shortened anatomical and functional survival in vitro (P < 0.01). The sulfhydryl donor, N-acetylcysteine (NAC; 10(-4) M), completely reversed both the positive and negative inotropic defects in Tat; increased GSH (P < 0.01) and GSH/GSSG (P < 0.01); reversed H(2)O(2) level (P < 0.05) and GPX1 activity (P < 0.05); and normalized the blunted inotropic response to Ca(2+) (P < 0.01). NAC (10(-7)) M normalized duration of contractile function from <40 min to >120 min (P < 0.01), with no effect on GSH and GSH/GSSG. NAC (10(-4) M) reverses cardiac myocyte dysfunction and markers of oxidative stress. NAC (10(-7) M) enhances myocyte function independent of changes in glutathione. Elucidating the molecular mechanisms involved in the GSH-dependent and GSH-independent salutary effects of NAC should identify novel therapeutic targets for myocardial proteinopathies recently appreciated in human cardiomyopathies.

Glutathione Dependent and Independent Salutary Effects of NAC on HIV Tat Proteinopathy

We have previously reported that cardiac myocyte‐specific expression of HIV‐Tat (Tat) results in a murine cardiomyopathy model. We now report that cardiac myocytes isolated from Tat exhibit decreased ATP (p&lt;0.01; n=8–9) and GSH levels (p&lt;0.01; n=8–9), decreased GSH/GSSG ratio (p&lt;0.01; n=8–9), increased H2O2 (p&lt;0.05; n=6) and catalase (p&lt;0.05; n=6), and GPX1 (glutathione peroxidase 1) activities (p&lt;0.05; n=6), blunted systolic function (%Fractional shortening, p&lt;0.01; n=15; +dl/dt p&lt;0.01; n=15) and diastolic function (‐dl/dt, p&lt;0.01; n=15), and inotropic responses to Ca++ (p&lt;0.01, n= 15) and shortened anatomical and functional survival in vitro (p&lt;0.01, n=15). Treatment of Tat cardiac myocytes with the sulfhydryl donor, N‐acetylcysteine (NAC; 10−4 M) reversed both the positive and negative inotropic defects in Tat; increased GSH (p&lt;0.01); GSH/GSSG (p&lt;0.01); reversed H2O2 level (p&lt;0.05) and GPX1 activity (p&lt;0.05); normalized the blunted inotropic response to Ca2+ (p&lt;0.01). In addition, NAC 10−7 M normalized duration of contractile function from &lt;40 min to &gt;120 min (p&lt;0.01), while having no affect on GSH, GSH/GSSG. Elucidating molecular mechanisms involved in the GSH‐dependent and GSH‐independent salutary effects of NAC could identify novel therapeutic targets to mitigate the pathogenic effects of proteinopthies in cardiomyopathies and heart failure.

Opposing Actions of Extracellular Signal-regulated Kinase (ERK) and Signal Transducer and Activator of Transcription 3 (STAT3) in Regulating Microtubule Stabilization during Cardiac Hypertrophy

Excessive proliferation and stabilization of the microtubule (MT) array in cardiac myocytes can accompany pathological cardiac hypertrophy, but the molecular control of these changes remains poorly characterized. In this study, we examined MT stabilization in two independent murine models of heart failure and revealed increases in the levels of post-translationally modified stable MTs, which were closely associated with STAT3 activation. To explore the molecular signaling events contributing to control of the cardiac MT network, we stimulated cardiac myocytes with an α-adrenergic agonist phenylephrine (PE), and observed increased tubulin content without changes in detyrosinated (glu-tubulin) stable MTs. In contrast, the hypertrophic interleukin-6 (IL6) family cytokines increased both the glu-tubulin content and glu-MT density. When we examined a role for ERK in regulating cardiac MTs, we showed that the MEK/ERK-inhibitor U0126 increased glu-MT density in either control cardiac myocytes or following exposure to hypertrophic agents. Conversely, expression of an activated MEK1 mutant reduced glu-tubulin levels. Thus, ERK signaling antagonizes stabilization of the cardiac MT array. In contrast, inhibiting either JAK2 with AG490, or STAT3 signaling with Stattic or siRNA knockdown, blocked cytokine-stimulated increases in glu-MT density. Furthermore, the expression of a constitutively active STAT3 mutant triggered increased glu-MT density in the absence of hypertrophic stimulation. Thus, STAT3 activation contributes substantially to cytokine-stimulated glu-MT changes. Taken together, our results highlight the opposing actions of STAT3 and ERK pathways in the regulation of MT changes associated with cardiac myocyte hypertrophy.

Anti-apoptotic potential of rosuvastatin pretreatment in murine model of cardiomyopathy

Background Apoptosis is a key pathologic feature in myocardial infarction and heart failure. Recent evidence suggests that statins may have beneficial effects on cardiovascular outcomes in patients with heart failure. The present study was planned to investigate the anti-apoptotic potential of rosuvastatin pretreatment in doxorubicin-induced cardiomyopathy. Methods Sixty male Wistar rats were randomly divided into six groups: Group-I (vehicle control group), Group-II (pathological Control group), Group-III (rosuvastatin 0.5 mg/kg), Group IV (rosuvastatin 2 mg/kg), Group-V (rosuvastatin 2 mg/kg per se) , and Group-VI (carvedilol 1 mg/kg). Myocardial apoptosis was detected by caspase-3 assay, DNA gel electrophoresis and Na +/K + ATPase estimation. The animals were evaluated for various biochemical parameters in serum followed by histopathological studies of heart tissue. Results Doxorubicin treated rats exhibited cardiac dysfunctions as indicated by an increase in systolic, diastolic, mean BP, heart rate and tail blood flow and volume and increased serum LDH, TC, TGs, LDL-C, VLDL-C levels and atherogenic indexes. A marked induction in caspase-3 and Na +-K + ATPase levels and DNA laddering as revealed by agarose gel electrophoresis was observed in rat myocardium of pathological group. Pretreatment with the test drug, rosuvastatin significantly reduced the increase in hemodynamic parameters, serum LDH, lipid profile and myocardial caspase-3, Na +-K + ATPase activity as compared to the pathogenic control group. Further, DNA ladder formation was attenuated by rosuvastatin treatment. Histopathological studies further confirm its myocardial salvaging effects. The results were comparable with carvedilol. Conclusions The study demonstrates the cardioprotective potential of rosuvastatin against doxorubicin-induced myocardial apoptosis.

Chronic Treatment with Clenbuterol Modulates Endothelial Progenitor Cells and Circulating Factors in a Murine Model of Cardiomyopathy

The purpose of this study was to determine the effects of chronic treatment with the beta 2 adrenergic receptor agonist clenbuterol on endothelial progenitor cells (EPC) in a well-characterized model of heart failure, the muscle LIM protein knockout (MLP(-/-)) mouse. MLP(-/-) mice were treated daily with clenbuterol (2 mg/kg) or saline subcutaneously for 6 weeks. Clenbuterol led to a 30% increase in CD31(+) cells in the bone marrow of MLP(-/-) heart failure mice (p < 0.004). Clenbuterol did not improve ejection fraction. Clenbuterol treatment in MLP(-/-) mice was associated with significant changes in the following circulating factors: tissue inhibitor of metalloproteinase-type 1, leukemia inhibitory factor 1, C-reactive protein, apolipoprotein A1, fibroblast growth factor 2, serum glutamic oxaloacetic transaminase, macrophage-derived chemokine, and monocyte chemoattractant protein-3. Clenbuterol treatment in the MLP(-/-) model of heart failure did not rescue heart function, yet did increase CD31(+) cells in the bone marrow. This is the first evidence that a beta 2 agonist increases EPC proliferation in the bone marrow in a preclinical model of heart failure.

Gene Therapy in Congestive Heart Failure

Although research and development of gene therapy have encountered several difficulties in recent years, they have also begun to show potential for the treatment of acquired and inherited human diseases. Gene therapy may have wide applicability for the treatment of numerous cardiovascular diseases, including congestive heart failure (CHF), ischemic heart disease, and cardiomyopathy. CHF remains a leading cause of death and morbidity in industrialized nations, and the number of patients with CHF continues to increase as the population ages. Pharmacological therapies control symptoms, reduce left ventricular (LV) dilation, improve function, and reduce mortality. These therapies control the disease but do not cure it. Currently, however, the growing knowledge of molecular mechanisms involved in myocardial dysfunction has allowed the identification of potential therapeutic targets designed with the aim of curing CHF. Several studies in animal models have provided hope that gene therapy may be one of the future strategies for the treatment of clinical CHF. In these studies, myocardial gene transfer has been used to target at least 3 different biological pathways that play a crucial role in the pathophysiology of CHF: Intracellular calcium signaling, β1-adrenergic receptor (β1-AR) signaling, and antiapoptotic signaling. The activity of sarcoplasmic reticulum Ca2+ ATPase (SERCA2a) is decreased in failing myocardium, and this results in abnormal Ca2+ handling.1 In an animal model of heart failure, Miyamoto et al2 showed that in vivo overexpression of SERCA2a, achieved by catheter injection of an adenovirus carrying the SERCA2a gene into the aortic root, restored systolic and diastolic function to normal levels. In another study, Muller et al3 analyzed the chronic overexpression of SERCA2a in both normal and pressure-overloaded hearts (induced by abdominal aortic banding of transgenic rats). Overexpression of SERCA2a resulted in a …