Speckle Tracking Based Strain Analysis Is Sensitive for Early Detection of Pathological Cardiac Hypertrophy

Cardiac hypertrophy is a key pathological process of many cardiac diseases. However, early detection of cardiac hypertrophy is difficult by the currently used non-invasive method and new approaches are in urgent need for efficient diagnosis of cardiac malfunction. Here we report that speckle tracking-based strain analysis is more sensitive than conventional echocardiography for early detection of pathological cardiac hypertrophy in the isoproterenol (ISO) mouse model. Pathological hypertrophy was induced by a single subcutaneous injection of ISO. Physiological cardiac hypertrophy was established by daily treadmill exercise for six weeks. Strain analysis, including radial strain (RS), radial strain rate (RSR) and longitudinal strain (LS), showed marked decrease as early as 3 days after ISO injection. Moreover, unlike the regional changes in cardiac infarction, strain analysis revealed global cardiac dysfunction that affects the entire heart in ISO-induced hypertrophy. In contrast, conventional echocardiography, only detected altered E/E’, an index reflecting cardiac diastolic function, at 7 days after ISO injection. No change was detected on fractional shortening (FS), E/A and E’/A’ at 3 days or 7 days after ISO injection. Interestingly, strain analysis revealed cardiac dysfunction only in ISO-induced pathological hypertrophy but not the physiological hypertrophy induced by exercise. Taken together, our study indicates that strain analysis offers a more sensitive approach for early detection of cardiac dysfunction than conventional echocardiography. Moreover, multiple strain readouts distinguish pathological cardiac hypertrophy from physiological hypertrophy.     

Overexpression of ornithine decarboxylase decreases ventricular systolic function during induction of cardiac hypertrophy

Ornithine decarboxylase (ODC), the first enzyme of polyamine metabolism, is rapidly upregulated in response to agents that induce a pathological cardiac hypertrophy. Transgenic mice overexpressing ODC in the heart (MHC-ODC mice) experience a much more dramatic left ventricular hypertrophy in response to β-adrenergic stimulation with isoproterenol (ISO) compared to wild-type (WT) controls. ISO also induced arginase activity in transgenic hearts but not in controls. The current work studies the cooperation between the cardiac polyamines and L-arginine (L-Arg) availability in MHC-ODC mice. Although ISO-induced hypertrophy is well-compensated, MHC-ODC mice administered L-Arg along with ISO showed a rapid onset of systolic dysfunction and died within 48 h. Myocytes isolated from MHC-ODC mice administered L-Arg/ISO exhibited reduced contractility and altered calcium transients, suggesting an alteration in [Ca(2+)] homeostasis, and abbreviated action potential duration, which may contribute to arrhythmogenesis. The already elevated levels of spermidine and spermine were not further altered in MHC-ODC hearts by L-Arg/ISO treatment, suggesting alternative L-Arg utilization pathways lead to dysregulation of intracellular calcium. MHC-ODC mice administered an arginase inhibitor (Nor-NOHA) along with ISO died almost as rapidly as L-Arg/ISO-treated mice, while the iNOS inhibitor S-methyl-isothiourea (SMT) was strongly protective against L-Arg/ISO. These results point to the induction of arginase as a protective response to β-adrenergic stimulation in the setting of high polyamines. Further, NO generated by exogenously supplied L-Arg may contribute to the lethal consequences of L-Arg/ISO treatment. Since considerable variations in human cardiac polyamine and L-Arg content are likely, it is possible that alterations in these factors may influence myocyte contractility.     

Alterations of beta-adrenergic signaling and cardiac hypertrophy in transgenic mice overexpressing TGF-beta(1)

Transforming growth factor-beta(1) (TGF-beta(1)) promotes or inhibits cell proliferation and induces fibrotic processes and extracellular matrix production in numerous cell types. Several cardiac diseases are associated with an increased expression of TGF-beta(1) mRNA, particularly during the transition from stable cardiac hypertrophy to heart failure. In vitro studies suggest a link between TGF-beta(1) signaling and the beta-adrenergic system. However, the in vivo effects of this growth factor on myocardial tissue have been poorly identified. In transgenic mice overexpressing TGF-beta(1) (TGF-beta), we investigated the in vivo effects on cardiac morphology, beta-adrenergic signaling, and contractile function. When compared with nontransgenic controls (NTG), TGF-beta mice revealed significant cardiac hypertrophy (heart weight, 164 +/- 7 vs. 130 +/- 3 mg, P < 0.01; heart weight-to-body weight ratio, 6.8 +/- 0.3 vs. 5.1 +/- 0.1 mg/g, P < 0.01), accompanied by interstitial fibrosis. These morphological changes correlated with an increased expression of hypertrophy-associated proteins such as atrial natriuretic factor (ANF). Furthermore, overexpression of TGF-beta(1) led to alterations of beta-adrenergic signaling as myocardial beta-adrenoceptor density increased from 7.3 +/- 0.3 to 11.2 +/- 1.1 fmol/mg protein (P < 0.05), whereas the expression of beta-adrenoceptor kinase-1 and inhibitory G proteins decreased by 56 +/- 9.7% and 58 +/- 7.6%, respectively (P < 0.05). As a consequence of altered beta-adrenergic signaling, hearts from TGF-beta showed enhanced contractile responsiveness to isoproterenol stimulation. In conclusion, we conclude that TGF-beta(1) induces cardiac hypertrophy and enhanced beta-adrenergic signaling in vivo. The morphological alterations are either induced by direct effects of TGF-beta(1) or may at least in part result from increased beta-adrenergic signaling, which may contribute to excessive catecholamine stimulation during the transition from compensated hypertrophy to heart failure.     

Alterations in apoptosis regulatory factors during hypertrophy and heart failure

Cardiac hypertrophy from pathological stimuli often proceeds to heart failure, whereas cardiac hypertrophy from physiological stimuli does not. In this study, physiological hypertrophy was created by a daily exercise regimen and pathological hypertrophy was created from a high-salt diet in Dahl salt-sensitive rats. The rats continued on a high-salt diet progressed to heart failure associated with an increased rate of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling-positive cardiomyocytes. We analyzed primary cultures of these hearts and found that only cardiomyocytes made hypertrophic by a pathological stimulus show increased sensitivity to apoptosis. Examination of the molecular changes associated with these distinct types of hypertrophy revealed changes in Bcl-2 family members and caspases favoring survival during physiological hypertrophy. However, in pathological hypertrophy, there were more diffuse proapoptotic changes, including changes in Fas, the Bcl-2 protein family, and caspases. Therefore, we speculate that this increased sensitivity to apoptotic stimulation along with proapoptotic changes in the apoptosis program may contribute to the development of heart failure seen in pathological cardiac hypertrophy.     

Plantago asiatica L. seeds extract protects against cardiomyocyte injury in isoproterenol- induced cardiac hypertrophy by inhibiting excessive autophagy and apoptosis in mice

BackgroundCardiac hypertrophy is the early stage of many heart diseases, such as coronary heart disease, hypertension, valvular dysfunction and cardiomyopathy. Cardiomyocyte autophagy and apoptosis play an important role in the process of cardiac hypertrophic response. Plantago asiatica L. seeds extract (PASE) is prepared from a traditional herbal medicine in Asia with tremendous pharmacological activities. However, whether PASE could relieve cardiac hypertrophy has not been elucidated. The present study is aimed to investigate the effect of PASE on cardiac hypertrophy and explore its potential underlying mechanism.MethodsCardiac hypertrophy was induced in C57BL/6 mice by subcutaneous injection of isoproterenol (ISO) for two weeks. Meanwhile, the mice were intraperitoneally injected with PASE at dosages of 20, 40 and 80 mg/kg/day. Cardiac hypertrophy was evaluated by echocardiographic examination, haematoxylin and eosin staining and quantitative real-time polymerase chain reaction. Expressions of proteins involved in autophagy and apoptosis such as Beclin1, p62, LC3II, Bax, Bcl-2 and Cleaved-caspase-3 were detected by western blot analysis. Western blot, transient transfection, acridine orange staining, TUNEL staining and autophagy inducer were used to observe the effect and explore the mechanism of PASE on cardiomyocyte and H9c2 cells with excessive autophagy and apoptosis induced by ISO.ResultsISO induction for two weeks disturbed the myocardial contractility and cardiac function of left ventricles of mice. PASE treated mice showed significantly improved cardiac function indexes, including EF, FS, SV and CO, compared with the ISO group. Treatment with PASE also decreased the heart weight/body weight ratio and cardiomyocyte size, and downregulated the mRNA and protein expressions of hypertrophic markers ANP, BNP, and β-MHC. Furthermore, the changes of autophagy and apoptosis markers, such as LC3II, Beclin1, p62, Bcl-2, Bax and Cleaved-caspase-3 induced by ISO were resumed by PASE treatment. Consistently, PASE demonstrated similar effects on ISO-induced H9c2 cells as it did in vivo. In addition, PASE could counteract the increased autophagy induced by the autophagy inducer, rapamycin.ConclusionPASE attenuated ISO-induced cardiac hypertrophy in mice by inhibiting excessive autophagy and apoptosis in cardiomyocytes. The novel findings may pave the way for the clinical usage of PASE for the prevention of heart diseases related with cardiac hypertrophy.     

LKB1IP promotes pathological cardiac hypertrophy by targeting PTEN/Akt signalling pathway

Pathological cardiac hypertrophy represents a leading cause of morbidity and mortality worldwide. Liver kinase B1 interacting protein 1 (LKB1IP) was identified as the binding protein of tumour suppressor LKB1. However, the role of LKB1IP in the development of pathological cardiac hypertrophy has not been explored. The aim of this study was to investigate the function of LKB1IP in cardiac hypertrophy in response to hypertrophic stimuli. We investigated the cardiac level of LKB1IP in samples from patients with heart failure and mice with cardiac hypertrophy induced by isoproterenol (ISO) or transverse aortic constriction (TAC). LKB1IP knockout mice were generated and challenged with ISO injection or TAC surgery. Cardiac function, hypertrophy and fibrosis were then examined. LKB1IP expression was significantly up-regulated on hypertrophic stimuli in both human and mouse cardiac samples. LKB1IP knockout markedly protected mouse hearts against ISO- or TAC-induced cardiac hypertrophy and fibrosis. LKB1IP overexpression aggravated ISO-induced cardiomyocyte hypertrophy, and its inhibition attenuated hypertrophy in vitro. Mechanistically, LKB1IP activated Akt signalling by directly targeting PTEN and then inhibiting its phosphatase activity. In conclusion, LKB1IP may be a potential target for pathological cardiac hypertrophy.     

Changes in short‐chain acyl‐coA dehydrogenase during rat cardiac development and stress

This study was designed to investigate the expression of short‐chain acyl‐CoA dehydrogenase (SCAD), a key enzyme of fatty acid β‐oxidation, during rat heart development and the difference of SCAD between pathological and physiological cardiac hypertrophy. The expression of SCAD was lowest in the foetal and neonatal heart, which had time‐dependent increase during normal heart development. In contrast, a significant decrease in SCAD expression was observed in different ages of spontaneously hypertensive rats (SHR). On the other hand, swim‐trained rats developed physiological cardiac hypertrophy, whereas SHR developed pathological cardiac hypertrophy. The two kinds of cardiac hypertrophy exhibited divergent SCAD changes in myocardial fatty acids utilization. In addition, the expression of SCAD was significantly decreased in pathological cardiomyocyte hypertrophy, however, increased in physiological cardiomyocyte hypertrophy. SCAD siRNA treatment triggered the pathological cardiomyocyte hypertrophy, which showed that the down‐regulation of SCAD expression may play an important role in pathological cardiac hypertrophy. The changes in peroxisome proliferator‐activated receptor α (PPARα) was accordant with that of SCAD. Moreover, the specific PPARα ligand fenofibrate treatment increased the expression of SCAD and inhibited pathological cardiac hypertrophy. Therefore, we speculate that the down‐regulated expression of SCAD in pathological cardiac hypertrophy may be responsible for ‘the recapitulation of foetal energy metabolism’. The deactivation of PPARα may result in the decrease in SCAD expression in pathological cardiac hypertrophy. Changes in SCAD are different in pathological and physiological cardiac hypertrophy, which may be used as the molecular markers of pathological and physiological cardiac hypertrophy.     

Overexpression of microRNA-99a Attenuates Cardiac Hypertrophy

Pathological cardiomyocyte hypertrophy is associated with significantly increased risk of heart failure, one of the leading medical causes of mortality worldwide. MicroRNAs are known to be involved in pathological cardiac remodeling. However, whether miR-99a participates in the signaling cascade leading to cardiac hypertrophy is unknown. To evaluate the role of miR-99a in cardiac hypertrophy, we assessed the expression of miR-99a in hypertrophic cardiomyocytes induced by isoprenaline (ISO)/angiotensin-II (Ang II) and in mice model of cardiac hypertrophy induced by transverse aortic constriction (TAC). Expression of miR-99a was evaluated in these hypertrophic cells and hearts. We also found that miR-99a expression was highly correlated with cardiac function of mice with heart failure (8 weeks after TAC surgery). Overexpression of miR-99a attenuated cardiac hypertrophy in TAC mice and cellular hypertrophy in stimuli treated cardiomyocytes through down-regulation of expression of mammalian target of rapamycin (mTOR). These results indicate that miR-99a negatively regulates physiological hypertrophy through mTOR signaling pathway, which may provide a new therapeutic approach for pressure-overload heart failure.     

Novel EGFR inhibitors attenuate cardiac hypertrophy induced by angiotensin II

Cardiac hypertrophy is an important risk factor for heart failure. Epidermal growth factor receptor (EGFR) has been found to play a role in the pathogenesis of various cardiovascular diseases. The aim of this current study was to examine the role of EGFR in angiotensin II (Ang II)‐induced cardiac hypertrophy and identify the underlying molecular mechanisms. In this study, we observed that both Ang II and EGF could increase the phospohorylation of EGFR and protein kinase B (AKT)/extracellular signal‐regulated kinase (ERK), and then induce cell hypertrophy in H9c2 cells. Both pharmacological inhibitors and genetic silencing significantly reduced Ang II‐induced EGFR signalling pathway activation, hypertrophic marker overexpression, and cell hypertrophy. In addition, our results showed that Ang II‐induced EGFR activation is mediated by c‐Src phosphorylation. In vivo, Ang II treatment significantly led to cardiac remodelling including cardiac hypertrophy, disorganization and fibrosis, accompanied by the activation of EGFR signalling pathway in the heart tissues, while all these molecular and pathological alterations were attenuated by the oral administration with EGFR inhibitors. In conclusion, the c‐Src‐dependent EGFR activation may play an important role in Ang II‐induced cardiac hypertrophy, and inhibition of EGFR by specific molecules may be an effective strategy for the treatment of Ang II‐associated cardiac diseases.     

Growth/differentiation factor 1 alleviates pressure overload-induced cardiac hypertrophy and dysfunction

Pathological cardiac hypertrophy is a major risk factor for developing heart failure, the leading cause of death in the world. Growth/differentiation factor 1 (GDF1), a transforming growth factor-β family member, is a regulator of cell growth and differentiation in both embryonic and adult tissues. Evidence from human and animal studies suggests that GDF1 may play an important role in cardiac physiology and pathology. However, a critical role for GDF1 in cardiac remodelling has not been investigated. Here, we performed gain-of-function and loss-of-function studies using cardiac-specific GDF1 knockout mice and transgenic mice to determine the role of GDF1 in pathological cardiac hypertrophy, which was induced by aortic banding (AB). The extent of cardiac hypertrophy was evaluated by echocardiographic, hemodynamic, pathological, and molecular analyses. Our results demonstrated that cardiac specific GDF1 overexpression in the heart markedly attenuated cardiac hypertrophy, fibrosis, and cardiac dysfunction, whereas loss of GDF1 in cardiomyocytes exaggerated the pathological cardiac hypertrophy and dysfunction in response to pressure overload. Mechanistically, we revealed that the cardioprotective effect of GDF1 on cardiac remodeling was associated with the inhibition of the MEK–ERK1/2 and Smad signaling cascades. Collectively, our data suggest that GDF1 plays a protective role in cardiac remodeling via the negative regulation of the MEK–ERK1/2 and Smad signaling pathways.     

Extracellular high‐mobility group box 1 mediates pressure overload‐induced cardiac hypertrophy and heart failure

Inflammation plays a key role in pressure overload‐induced cardiac hypertrophy and heart failure, but the mechanisms have not been fully elucidated. High‐mobility group box 1 (HMGB1), which is increased in myocardium under pressure overload, may be involved in pressure overload‐induced cardiac injury. The objectives of this study are to determine the role of HMGB1 in cardiac hypertrophy and cardiac dysfunction under pressure overload. Pressure overload was imposed on the heart of male wild‐type mice by transverse aortic constriction (TAC), while recombinant HMGB1, HMGB1 box A (a competitive antagonist of HMGB1) or PBS was injected into the LV wall. Moreover, cardiac myocytes were cultured and given sustained mechanical stress. Transthoracic echocardiography was performed after the operation and sections for histological analyses were generated from paraffin‐embedded hearts. Relevant proteins and genes were detected. Cardiac HMGB1 expression was increased after TAC, which was accompanied by its translocation from nucleus to both cytoplasm and intercellular space. Exogenous HMGB1 aggravated TAC‐induced cardiac hypertrophy and cardiac dysfunction, as demonstrated by echocardiographic analyses, histological analyses and foetal cardiac genes detection. Nevertheless, the aforementioned pathological change induced by TAC could partially be reversed by HMGB1 inhibition. Consistent with the in vivo observations, mechanical stress evoked the release and synthesis of HMGB1 in cultured cardiac myocytes. This study indicates that the activated and up‐regulated HMGB1 in myocardium, which might partially be derived from cardiac myocytes under pressure overload, may be of crucial importance in pressure overload‐induced cardiac hypertrophy and cardiac dysfunction.     

RASSF1A: A potential novel therapeutic target against cardiac hypertrophy

Cardiac hypertrophy is a key risk factor for chronic heart failure. Current treatments predominantly focus on both reducing the peripheral vascular resistance and activating nerve-humoral system. However, these efforts can't reverse cardiac hypertrophy fundamentally. Ras association domain family 1 isoform A (RASSF1A) is a regulatory tumor suppressor whose inactivation by inappropriate promoter methylation has been implicated in the development of many human cancers. Recently, there have been a number of studies investigating the roles of RASSF1A in the pathophysiology of cardiac hypertrophy. In this review, we focus on the present progresses of cardiac RASSF1A under physiological and pathological conditions, trying to systematically elucidate how the RASSF1A-mediated signal pathways contribute to the maintenance of normal cardiac myocyte structure and function and lead to the regression of pathological cardiac hypertrophy. These pathways exert multiple functions such as regulating cardiac contractility, physiologically increasing stability of microtubule, preventing cardiac dysfunction, attenuating interstitial fibrosis and mediating cell apoptosis. These specific roles are highly relevant with cardiac hemodynamics and therapeutic strategies, indicating RASSF1A may have the potential to reverse pathological cardiac hypertrophy thus prevent heart failure fundamentally.     

Alterations of the Z-band in cardiac and skeletal muscles.

The heart muscle may react to various hypoxic damaging effects (e.g.N2, CO, haemorrhagic shock, electroshock) by identical responses similarly as in the case of skeletal muscle damages. One of the early manifestations of the process is the alteration of the Z-band, which is considered pathological. The alterations of the Z-band region precede the changes of the mitochondrial and sacrotubular systems and might form the morphological basis of functional changes induced by hypoxic effects. The alterations observed are differentiated from the hypertrophy of the Z-band. In the development of the alterations of the Z-band the role of other factors (e.g. calcium metabolism, sacroplasmatic membrane changes) is emphasized.     

Regional Cardiac Dysfunction and Dyssynchrony in a Murine Model of Afterload Stress

Small animal models of afterload stress have contributed much to our present understanding of the progression from hypertension to heart failure. High-sensitivity methods for phenotyping cardiac function in vivo, particular in the setting of compensated cardiac hypertrophy, may add new information regarding alterations in cardiac performance that can occur even during the earliest stages of exposure to pressure overload. We have developed an echocardiographic analytical method, based on speckle-tracking-based strain analyses, and used this tool to rapidly phenotype cardiac changes resulting from afterload stress in a small animal model. Adult mice were subjected to ascending aortic constriction, with and without subsequent reversal of the pressure gradient. In this model of compensated hypertrophic cardiac remodeling, conventional echocardiographic measurements did not detect changes in left ventricular (LV) function at the early time points examined. Strain analyses, however, revealed a decrement in basal longitudinal myofiber shortening that was induced by aortic constriction and improved following relief of the pressure gradient. Furthermore, we observed that pressure overload resulted in LV segmental dyssynchrony that was attenuated with return of the afterload to baseline levels. Herein, we describe the use of echocardiographic strain analyses for cardiac phenotyping in a mouse model of pressure overload. This method provides evidence of dyssynchrony and regional myocardial dysfunction that occurs early with compensatory hypertrophy, and improves following relief of aortic constriction. Importantly, these findings illustrate the utility of a rapid, non-invasive method for characterizing early cardiac dysfunction, not detectable by conventional echocardiography, following afterload stress.