echocardiographic-characterization-left-ventricular-structure

Echocardiographic Characterization of Left Ventricular Structure, Function, and Coronary Flow in Neonate Mice

Echocardiography is a non-invasive procedure that enables the evaluation of structural and functional parameters in animal models of cardiovascular ...

University of Illinois at Chicago

Discrepancies exist regarding potential sex differences in the effects of ethanol on the myocardium. Therefore, the aims of this study were to determine if long-term ethanol consumption was associated with the development of a dilated cardiomyopathy (CM) in female rats and, second, to determine if the absence of ovarian hormones modulated this effect. Adult male and female (n=6-8/group) sham-operated and ovariectomized (OVX) Sprague-Dawley rats received the Lieber DeCarli ethanol-containing (8% vol/vol) or control liquid diet for 8 months. All ethanol groups showed echocardiographic evidence of a cardiomyopathy; however, more significant ethanol-elicited differences were found in the male ethanol group than in either the female or female OVX groups. In addition, the male ethanol group had significant reductions in in vivo measures of contractility, such as the maximum derivative of change in systolic pressure and preload recruitable stroke work. Sex differences were also apparent in the pattern and degree of posterior and septal wall thickness changes, in that the male ethanol group had more posterior and septal wall thinning. In conclusion, similar to male rats long-term ethanol consumption in gonad-intact and OVX female rats is associated with the development of a dilated cardiomyopathy.

Long-term effects of alcohol consumption in male and female rats.

To determine the effects of cigarette smoke (CS) exposure on the expression/activation of mitogen-activated protein kinases (MAPKs) (extracellular signal-regulated kinase [ERK1/2], p38-kinase [p38] and c-Jun NH2-terminal protein kinase [JNK]), norepinephrine (NE) levels and myocardial structure and function.

Cigarette smoke-induced left ventricular remodelling is associated with activation of mitogen-activated protein kinases.

The effects of nicotine (NIC) on normal hearts are fairly well established, yet its effects on hearts displaying familial hypertrophic cardiomyopathy have not been tested. We studied both the acute and chronic effects of NIC on a transgenic (TG) mouse model of FHC caused by a mutation in α-tropomyosin (Tm; i.e., α-Tm D175N TG, or Tm175). For acute effects, intravenously injected NIC increased heart rate, left ventricular (LV) pressure, and the maximal rate of LV pressure increase (+dP/dt) in non-TG (NTG) and Tm175 mice; however, Tm175 showed a significantly smaller increase in the maximal rate of LV pressure decrease (-dP/dt) compared with NTGs. Western blots revealed phosphorylation of phospholamban Ser16 and Thr17 residue increased in NTG mice following NIC injection but not in Tm175 mice. In contrast, phosphorylation of troponin I at serine residues 23 and 24 increased equally in both NTG and Tm175. Thus the attenuated increase in relaxation in Tm175 mice following acute NIC appears to result primarily from attenuated phospholamban phosphorylation. Chronic NIC administration (equivalent to smoking 2 packs of cigarettes/day for 4 mo) also increased +dP/dt in NTG and Tm175 mice compared with chronic saline. However, chronic NIC had little effect on heart rate, LV pressure, -dP/dt, LV wall and chamber dimensions, or collagen content for either group of mice.

Effects of nicotine administration in a mouse model of familial hypertrophic cardiomyopathy, α-tropomyosin D175N.

We have recently shown that a temporary increase in sarcoplasmic reticulum (SR) cycling via adenovirus-mediated overexpression of sarcoplasmic reticulum ATPase (SERCA2) transiently improves relaxation and delays hypertrophic remodeling in a familial hypertrophic cardiomyopathy (FHC) caused by a mutation in the thin filament protein, tropomyosin (i.e., α-TmE180G or Tm180). In this study, we sought to permanently alter calcium fluxes via phospholamban (PLN) gene deletion in Tm180 mice in order to sustain long-term improvements in cardiac function and adverse cardiac remodeling/hypertrophy. While similar work has been done in FHCs resulting from mutations in thick myofilament proteins, no one has studied these effects in an FHC resulting from a thin filament protein mutation. Tm180 transgenic (TG) mice were crossbred with PLN knockout (KO) mice and four groups were studied in parallel: 1) non-TG (NTG), 2) Tm180, 3) PLNKO/NTG and 4) PLNKO/Tm180. Tm180 mice exhibit increased heart weight/body weight and hypertrophic gene markers compared to NTG mice, but levels in PLNKO/Tm180 mice were similar to NTG. Tm180 mice also displayed altered function as assessed via in situ pressure-volume analysis and echocardiography at 3-6 months and one year; however, altered function in Tm180 mice was rescued back to NTG levels in PLNKO/Tm180 mice. Collagen deposition, as assessed by Picrosirius Red staining, was increased in Tm180 mice but was similar in NTG and in PLNKO/Tm180 mice. Extracellular signal-regulated kinase (ERK1/2) phosphorylation increased in Tm180 mice while levels in PLNKO/Tm180 mice were similar to NTGs. The present study shows that by modulating SR calcium cycling, we were able to rescue many of the deleterious aspects of FHC caused by a mutation in the thin filament protein, Tm.

Long-term rescue of a familial hypertrophic cardiomyopathy caused by a mutation in the thin filament protein, tropomyosin, via modulation of a calcium cycling protein.

Excessive alcohol use in the form of binge drinking is associated with many adverse medical outcomes. Using an animal model, the primary objective of this study was to determine the effects of repeated episodes of binge drinking on myocardial structure, blood pressure (BP) and activation of mitogen-activated protein kinases (MAPKs). The effects of carvedilol, a beta-adrenergic blocker, were also examined in this animal model of binge drinking.

Cardiovascular responses and differential changes in mitogen-activated protein kinases following repeated episodes of binge drinking.

During development and differentiation, cell type-specific chromatin configurations are set up to facilitate cell type-specific gene expression. Defects in the establishment or the maintenance of the correct chromatin configuration have been associated with diseases ranging from leukemia to muscular dystrophy. The heart expresses many chromatin factors, and we are only beginning to understand their roles in heart development and function. We have previously shown that the chromatin regulator Asxl2 is highly expressed in the murine heart both during development and adulthood. In the absence of Asxl2, there is a significant reduction in trimethylation of histone H3 lysine 27 (H3K27), a histone mark associated with lineage-specific silencing of developmental genes. Here we present evidence that Asxl2 is required for the long-term maintenance of ventricular function and for the maintenance of normal cardiac gene expression. Asxl2(-/-) hearts displayed progressive deterioration of ventricular function. By 10 months of age, there was ~37% reduction in fractional shortening in Asxl2(-/-) hearts compared to wild-type. Analysis of the expression of myofibril proteins suggests that Asxl2 is required for the repression of β-MHC. Asxl2(-/-) hearts did not exhibit hypertrophy, suggesting that the de-repression of β-MHC was not the result of hypertrophic response. Instead, Asxl2 and the histone methyltansferase Ezh2 co-localize to β-MHC promoter, suggesting that Asxl2 directly represses β-MHC. Interrogation of the CardioGenomics database revealed that ASXL2 is down-regulated in the hearts of patients with ischemic or idiopathic dilated cardiomyopathy. We propose that chromatin factors like Asxl2 function in the adult heart to regulate cell type- and stage-specific patterns of gene expression, and the disruption of such regulation may be involved in the etiology and/or development of certain forms of human heart disease.

Maintenance of adult cardiac function requires the chromatin factor Asxl2.

α-Tropomyosin (α-TM) has a conserved, charged Asp-137 residue located in the hydrophobic core of its coiled-coil structure, which is unusual in that the residue is found at a position typically occupied by a hydrophobic residue. Asp-137 is thought to destabilize the coiled-coil and so impart structural flexibility to the molecule, which is believed to be crucial for its function in the heart. A previous in vitro study indicated that the conversion of Asp-137 to a more typical canonical Leu alters flexibility of TM and affects its in vitro regulatory functions. However, the physiological importance of the residue Asp-137 and altered TM flexibility is unknown. In this study, we further analyzed structural properties of the α-TM-D137L variant and addressed the physiological importance of TM flexibility in cardiac function in studies with a novel transgenic mouse model expressing α-TM-D137L in the heart. Our NMR spectroscopy data indicated that the presence of D137L introduced long range rearrangements in TM structure. Differential scanning calorimetry measurements demonstrated that α-TM-D137L has higher thermal stability compared with α-TM, which correlated with decreased flexibility. Hearts of transgenic mice expressing α-TM-D137L showed systolic and diastolic dysfunction with decreased myofilament Ca(2+) sensitivity and cardiomyocyte contractility without changes in intracellular Ca(2+) transients or post-translational modifications of major myofilament proteins. We conclude that conversion of the highly conserved Asp-137 to Leu results in loss of flexibility of TM that is important for its regulatory functions in mouse hearts. Thus, our results provide insight into the link between flexibility of TM and its function in ejecting hearts.

Conserved Asp-137 is important for both structure and regulatory functions of cardiac α-tropomyosin (α-TM) in a novel transgenic mouse model expressing α-TM-D137L.

Although ceramide accumulation in the heart is considered a major factor in promoting apoptosis and cardiac disorders, including heart failure, lipotoxicity and ischemia-reperfusion injury, little is known about ceramide's role in mediating changes in contractility. In the present study, we measured the functional consequences of acute exposure of isolated field-stimulated adult rat cardiomyocytes to C6-ceramide. Exogenous ceramide treatment depressed the peak amplitude and the maximal velocity of shortening without altering intracellular calcium levels or kinetics. The inactive ceramide analog C6-dihydroceramide had no effect on myocyte shortening or [Ca(2+)]i transients. Experiments testing a potential role for C6-ceramide-mediated effects on activation of protein kinase C (PKC) demonstrated evidence for signaling through the calcium-independent isoform, PKCε. We employed 2-dimensional electrophoresis and anti-phospho-peptide antibodies to test whether treatment of the cardiomyocytes with C6-ceramide altered myocyte shortening via PKC-dependent phosphorylation of myofilament proteins. Compared to controls, myocytes treated with ceramide exhibited increased phosphorylation of myosin binding protein-C (cMyBP-C), specifically at Ser273 and Ser302, and troponin I (cTnI) at sites apart from Ser23/24, which could be attenuated with PKC inhibition. We conclude that the altered myofilament response to calcium resulting from multiple sites of PKC-dependent phosphorylation contributes to contractile dysfunction that is associated with cardiac diseases in which elevations in ceramides are present. 

Ceramide-mediated depression in cardiomyocyte contractility through PKC activation and modulation of myofilament protein phosphorylation.

Despite its known cardiovascular benefits, the intracellular signalling mechanisms underlying physiological cardiac growth remain poorly understood. Therefore, the purpose of this study was to investigate a novel role of p21-activated kinase-1 (Pak1) in the regulation of exercise-induced cardiac hypertrophy.

Knockout of p21-activated kinase-1 attenuates exercise-induced cardiac remodelling through altered calcineurin signalling.

Hypertrophic cardiomyopathy (HCM) is a genetic cardiovascular disorder, primarily involving mutations in sarcomeric proteins. HCM patients present with hypertrophy, diastolic dysfunction, and fibrosis, but there is no specific treatment. The sphingosine-1-phosphate receptor modulator, FTY720/fingolimod, is approved for treatment of multiple sclerosis. We hypothesize that modulation of the sphingosine-1-phosphate receptor by FTY720 would be of therapeutic benefit in sarcomere-linked HCM.

Sphingosine-1-Phosphate Receptor Modulator, FTY720, Improves Diastolic Dysfunction and Partially Reverses Atrial Remodeling in a Tm-E180G Mouse Model Linked to Hypertrophic Cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is a genetic disorder caused by mutations in different genes mainly encoding myofilament proteins and therefore called a "disease of the sarcomere." Despite the discovery of sarcomere protein mutations linked to HCM almost 30 years ago, the cellular mechanisms responsible for the development of this disease are not completely understood and likely vary among different mutations. Moreover, despite many efforts to develop effective treatments for HCM, these have largely been unsuccessful, and more studies are needed to better understand the cellular mechanisms of the disease. In experiments reported here, we investigated a mouse model expressing the mutant cTnT-R92Q, which is linked to HCM and induces an increase in myofilament Ca sensitivity and diastolic dysfunction. We found that early correction of the diastolic dysfunction by phospholamban knockout (PLNKO) was able to prevent the development of the HCM phenotype in troponin T (TnT)-R92Q transgenic (TG) mice. Four groups of mice in FVB/N background were generated and used for the experiments: (1) non-transgenic (NTG)/PLN mice, which express wild-type TnT and normal level of PLN; (2) NTG/PLNKO mice, which express wild-type TnT and no PLN; (3) TG/PLN mice, which express TnT-R92Q and normal level of PLN; (4) TG/PLNKO mice, which express TnT-R92Q and no PLN. Cardiac function was determined using both standard echocardiographic parameters and speckle tracking strain measurements. We found that both atrial morphology and diastolic function were altered in TG/PLN mice but normal in TG/PLNKO mice. Histological analysis showed a disarray of myocytes and increased collagen deposition only in TG/PLN hearts. We also observed increased Ca/calmodulin-dependent protein kinase II (CaMKII) phosphorylation only in TG/PLN hearts but not in TG/PLNKO hearts. The rescue of the HCM phenotype was not associated with differences in myofilament Ca sensitivity between TG/PLN and TG/PLNKO mice. Moreover, compared to standard systolic echo parameters, such as ejection fraction (EF), speckle strain measurements provided a more sensitive approach to detect early systolic dysfunction in TG/PLN mice. In summary, our results indicate that targeting diastolic dysfunction through altering Ca fluxes with no change in myofilament response to Ca was able to prevent the development of the HCM phenotype and should be considered as a potential additional treatment for HCM patients.

Modifications of Sarcoplasmic Reticulum Function Prevent Progression of Sarcomere-Linked Hypertrophic Cardiomyopathy Despite a Persistent Increase in Myofilament Calcium Response.

Experiments reported here tested the hypothesis that β-arrestin-2 is an important element in the preservation of cardiac function during aging. We tested this hypothesis by aging β-arrestin-2 knock-out (KO) mice, and wild-type equivalent (WT) to 12-16months. We developed the rationale for these experiments on the basis that angiotensin II (ang II) signaling at ang II receptor type 1 (AT1R), which is a G-protein coupled receptor (GPCR) promotes both G-protein signaling as well as β-arrestin-2 signaling. β-arrestin-2 participates in GPCR desensitization, internalization, but also acts as a scaffold for adaptive signal transduction that may occur independently or in parallel to G-protein signaling. We have previously reported that biased ligands acting at the AT1R promote β-arrestin-2 signaling increasing cardiac contractility and reducing maladaptations in a mouse model of dilated cardiomyopathy. Although there is evidence that ang II induces maladaptive senescence in the cardiovascular system, a role for β-arrestin-2 signaling has not been studied in aging. By echocardiography, we found that compared to controls aged KO mice exhibited enlarged left atria and left ventricular diameters as well as depressed contractility parameters with preserved ejection fraction. Aged KO also exhibited depressed relaxation parameters when compared to WT controls at the same age. Moreover, cardiac dysfunction in aged KO mice was correlated with alterations in the phosphorylation of myofilament proteins, such as cardiac myosin binding protein-C, and myosin regulatory light chain. Our evidence provides novel insights into a role for β-arrestin-2 as an important signaling mechanism that preserves cardiac function during aging.

Article	Year
Long-term effects of alcohol consumption in male and female rats. Cardiovascular toxicology\| PubMed ID: 17990129	2007
Cigarette smoke-induced left ventricular remodelling is associated with activation of mitogen-activated protein kinases. European journal of heart failure\| PubMed ID: 18815071	2008
Effects of nicotine administration in a mouse model of familial hypertrophic cardiomyopathy, α-tropomyosin D175N. American journal of physiology. Heart and circulatory physiology\| PubMed ID: 21743000	2011
Long-term rescue of a familial hypertrophic cardiomyopathy caused by a mutation in the thin filament protein, tropomyosin, via modulation of a calcium cycling protein. Journal of molecular and cellular cardiology\| PubMed ID: 21840315	2011
Cardiovascular responses and differential changes in mitogen-activated protein kinases following repeated episodes of binge drinking. Alcohol and alcoholism (Oxford, Oxfordshire)\| PubMed ID: 22878590	2013
Maintenance of adult cardiac function requires the chromatin factor Asxl2. Journal of molecular and cellular cardiology\| PubMed ID: 23046516	2012
Conserved Asp-137 is important for both structure and regulatory functions of cardiac α-tropomyosin (α-TM) in a novel transgenic mouse model expressing α-TM-D137L. The Journal of biological chemistry\| PubMed ID: 23609439	2013
Ceramide-mediated depression in cardiomyocyte contractility through PKC activation and modulation of myofilament protein phosphorylation. Basic research in cardiology\| PubMed ID: 25280528	2014
Knockout of p21-activated kinase-1 attenuates exercise-induced cardiac remodelling through altered calcineurin signalling. Cardiovascular research\| PubMed ID: 26464331	2015
Sphingosine-1-Phosphate Receptor Modulator, FTY720, Improves Diastolic Dysfunction and Partially Reverses Atrial Remodeling in a Tm-E180G Mouse Model Linked to Hypertrophic Cardiomyopathy. Circulation. Heart failure\| PubMed ID: 31684756	2019
Modifications of Sarcoplasmic Reticulum Function Prevent Progression of Sarcomere-Linked Hypertrophic Cardiomyopathy Despite a Persistent Increase in Myofilament Calcium Response. Frontiers in physiology\| PubMed ID: 32210830	2020
B-arrestin-2 Signaling Is Important to Preserve Cardiac Function During Aging. Frontiers in physiology\| PubMed ID: 34512376	2021

Shamim A. K. Chowdhury

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