Maladaptive Cardiac Hypertrophy Research Articles

Role of Vasoactive Hormone-Induced Signal Transduction in Cardiac Hypertrophy and Heart Failure.

Heart failure is the common concluding pathway for a majority of cardiovascular diseases and is associated with cardiac dysfunction. Since heart failure is invariably preceded by adaptive or maladaptive cardiac hypertrophy, several biochemical mechanisms have been proposed to explain the development of cardiac hypertrophy and progression to heart failure. One of these includes the activation of different neuroendocrine systems for elevating the circulating levels of different vasoactive hormones such as catecholamines, angiotensin II, vasopressin, serotonin and endothelins. All these hormones are released in the circulation and stimulate different signal transduction systems by acting on their respective receptors on the cell membrane to promote protein synthesis in cardiomyocytes and induce cardiac hypertrophy. The elevated levels of these vasoactive hormones induce hemodynamic overload, increase ventricular wall tension, increase protein synthesis and the occurrence of cardiac remodeling. In addition, there occurs an increase in proinflammatory cytokines and collagen synthesis for the induction of myocardial fibrosis and the transition of adaptive to maladaptive hypertrophy. The prolonged exposure of the hypertrophied heart to these vasoactive hormones has been reported to result in the oxidation of catecholamines and serotonin via monoamine oxidase as well as the activation of NADPH oxidase via angiotensin II and endothelins to promote oxidative stress. The development of oxidative stress produces subcellular defects, Ca2+-handling abnormalities, mitochondrial Ca2+-overload and cardiac dysfunction by activating different proteases and depressing cardiac gene expression, in addition to destabilizing the extracellular matrix upon activating some metalloproteinases. These observations support the view that elevated levels of various vasoactive hormones, by producing hemodynamic overload and activating their respective receptor-mediated signal transduction mechanisms, induce cardiac hypertrophy. Furthermore, the occurrence of oxidative stress due to the prolonged exposure of the hypertrophied heart to these hormones plays a critical role in the progression of heart failure.

Cardiac hypertrophy that affects hyperthyroidism occurs independently of the NLRP3 inflammasome.

Cardiac hypertrophy (CH) is an adaptive response to maintain cardiac function; however, persistent stress responses lead to contractile dysfunction and heart failure. Although inflammation is involved in these processes, the mechanisms that control cardiac inflammation and hypertrophy still need to be clarified. The NLRP3 inflammasome is a cytosolic multiprotein complex that mediates IL-1β production. The priming step of NLRP3 is essential for increasing the expression of its components and occurs following NF-κB activation. Hyperthyroidism triggers CH, which can progress to maladaptive CH and even heart failure. We have shown in a previous study that thyroid hormone (TH)-induced CH is linked to the upregulation of S100A8, leading to NF-κB activation. Therefore, we aimed to investigate whether the NLRP3 inflammasome is involved in TH-induced CH and its potential role in CH pathophysiology. Hyperthyroidism was induced in NLRP3 knockout (NLRP3-KO), Caspase-1-KO and Wild Type (WT) male mice of the C57Bl/6J strain, aged 8-12 weeks, by triiodothyronine (7μg/100g BW, i.p.) administered daily for 14 days. Morphological and cardiac functional analysis besides molecular assays showed, for the first time, that TH-induced CH is accompanied by reduced NLRP3 expression in the heart and that it occurs independently of the NLRP3 inflammasome and caspase 1-related pathways. However, NLRP3 is important for the maintenance of basal cardiac function since NLRP3-KO mice had impaired diastolic function and reduced heart rate, ejection fraction, and fractional shortening compared with WT mice.

The redox-active defensive Selenoprotein T as a novel stress sensor protein playing a key role in the pathophysiology of heart failure

Maladaptive cardiac hypertrophy contributes to the development of heart failure (HF). The oxidoreductase Selenoprotein T (SELENOT) emerged as a key regulator during rat cardiogenesis and acute cardiac protection. However, its action in chronic settings of cardiac dysfunction is not understood. Here, we investigated the role of SELENOT in the pathophysiology of HF: (i) by designing a small peptide (PSELT), recapitulating SELENOT activity via the redox site, and assessed its beneficial action in a preclinical model of HF [aged spontaneously hypertensive heart failure (SHHF) rats] and against isoproterenol (ISO)-induced hypertrophy in rat ventricular H9c2 and adult human AC16 cardiomyocytes; (ii) by evaluating the SELENOT intra-cardiomyocyte production and secretion under hypertrophied stimulation. Results showed that PSELT attenuated systemic inflammation, lipopolysaccharide (LPS)-induced macrophage M1 polarization, myocardial injury, and the severe ultrastructural alterations, while counteracting key mediators of cardiac fibrosis, aging, and DNA damage and restoring desmin downregulation and SELENOT upregulation in the failing hearts. In the hemodynamic assessment, PSELT improved the contractile impairment at baseline and following ischemia/reperfusion injury, and reduced infarct size in normal and failing hearts. At cellular level, PSELT counteracted ISO-mediated hypertrophy and ultrastructural alterations through its redox motif, while mitigating ISO-triggered SELENOT intracellular production and secretion, a phenomenon that presumably reflects the extent of cell damage. Altogether, these results indicate that SELENOT could represent a novel sensor of hypertrophied cardiomyocytes and a potential PSELT-based new therapeutic approach in myocardial hypertrophy and HF.Graphical

Behavior of Hypertrophied Right Ventricle during the Development of Left Ventricular Failure Due to Myocardial Infarction.

In order to determine the behavior of the right ventricle, we have reviewed the existing literature in the area of cardiac remodeling, signal transduction pathways, subcellular mechanisms, β-adrenoreceptor-adenylyl cyclase system and myocardial catecholamine content during the development of left ventricular failure due to myocardial infarction. The right ventricle exhibited adaptive cardiac hypertrophy due to increases in different signal transduction pathways involving the activation of protein kinase C, phospholipase C and protein kinase A systems by elevated levels of vasoactive hormones such as catecholamines and angiotensin II in the circulation at early and moderate stages of heart failure. An increase in the sarcoplasmic reticulum Ca2+ transport without any changes in myofibrillar Ca2+-stimulated ATPase was observed in the right ventricle at early and moderate stages of heart failure. On the other hand, the right ventricle showed maladaptive cardiac hypertrophy at the severe stages of heart failure due to myocardial infarction. The upregulation and downregulation of β-adrenoreceptor-mediated signal transduction pathways were observed in the right ventricle at moderate and late stages of heart failure, respectively. The catalytic activity of adenylate cyclase, as well as the regulation of this enzyme by Gs proteins, were seen to be augmented in the hypertrophied right ventricle at early, moderate and severe stages of heart failure. Furthermore, catecholamine stores and catecholamine uptake in the right ventricle were also affected as a consequence of changes in the sympathetic nervous system at different stages of heart failure. It is suggested that the hypertrophied right ventricle may serve as a compensatory mechanism to the left ventricle during the development of early and moderate stages of heart failure.

Cardiac recovery from pressure overload is not altered by thyroid hormone status in old mice.

Thyroid hormones (THs) are known to have various effects on the cardiovascular system. However, the impact of TH levels on preexisting cardiac diseases is still unclear. Pressure overload due to arterial hypertension or aortic stenosis and aging are major risk factors for the development of structural and functional abnormalities and subsequent heart failure. Here, we assessed the sensitivity to altered TH levels in aged mice with maladaptive cardiac hypertrophy and cardiac dysfunction induced by transverse aortic constriction (TAC). Mice at the age of 12 months underwent TAC and received T4 or anti-thyroid medication in drinking water over the course of 4 weeks after induction of left ventricular pressure overload. T4 excess or deprivation in older mice had no or only very little impact on cardiac function (fractional shortening), cardiac remodeling (cardiac wall thickness, heart weight, cardiomyocyte size, apoptosis, and interstitial fibrosis), and mortality. This is surprising because T4 excess or deprivation had significantly changed the outcome after TAC in young 8-week-old mice. Comparing the gene expression of deiodinases (Dio) 2 and 3 and TH receptor alpha (TRα) 1 and the dominant-negative acting isoform TRα2 between young and aged mice revealed that aged mice exhibited a higher expression of TRα2 and Dio3, while expression of Dio2 was reduced compared with young mice. These changes in Dio2 and 3 expressions might lead to reduced TH availability in the hearts of 12-month-old mice accompanied by reduced TRα action due to higher TRα2. In summary, our study shows that low and high TH availability have little impact on cardiac function and remodeling in older mice with preexisting pressure-induced cardiac damage. This observation seems to be the result of an altered expression of deiodinases and TRα isoforms, thus suggesting that even though cardiovascular risk is increasing with age, the response to TH stress may be dampened in certain conditions.

TIGAR Deficiency Blunts Angiotensin-II-Induced Cardiac Hypertrophy in Mice.

Hypertension is the key contributor to pathological cardiac hypertrophy. Growing evidence indicates that glucose metabolism plays an essential role in cardiac hypertrophy. TP53-induced glycolysis and apoptosis regulator (TIGAR) has been shown to regulate glucose metabolism in pressure overload-induced cardiac remodeling. In the present study, we investigated the role of TIGAR in cardiac remodeling during Angiotensin II (Ang-II)-induced hypertension. Wild-type (WT) and TIGAR knockout (KO) mice were infused with Angiotensin-II (Ang-II, 1 µg/kg/min) via mini-pump for four weeks. The blood pressure was similar between the WT and TIGAR KO mice. The Ang-II infusion resulted in a similar reduction of systolic function in both groups, as evidenced by the comparable decrease in LV ejection fraction and fractional shortening. The Ang-II infusion also increased the isovolumic relaxation time and myocardial performance index to the same extent in WT and TIGAR KO mice, suggesting the development of similar diastolic dysfunction. However, the knockout of TIGAR significantly attenuated hypertension-induced cardiac hypertrophy. This was associated with higher levels of fructose 2,6-bisphosphate, PFK-1, and Glut-4 in the TIGAR KO mice. Our present study suggests that TIGAR is involved in the control of glucose metabolism and glucose transporters by Ang-II and that knockout of TIGAR attenuates the development of maladaptive cardiac hypertrophy.

Inactivation of catalytic and non-catalytic PI3-kinase gamma function enhances hypoxia-induced pulmonary hypertension in mice

Abstract Rationale: Pulmonary hypertension (PH) is a pulmonary vascular disease characterised by chronically elevated pulmonary arterial mean pressure, increased pulmonary vascular resistance and right ventricular (RV) dysfunction and hypertrophy. The pathogenesis is characterised by vasoconstriction of pulmonary vessels and pulmonary vascular remodelling. PI 3-kinase γ (PI3Kγ) is activated via G protein-coupled receptors and abundantly expressed in all cell types involved in the pathogenesis of PH including leukocytes, cardiomyocytes, and endothelial cells (EC). The catalytic function of PI3Kγ is involved in numerous processes that are important for both vascular remodelling and maladaptive cardiac hypertrophy, including leukocyte recruitment and EC proliferation and survival. On the other hand, its non-catalytic function affects cyclic adenosine monophosphate (cAMP) production and thus cardiac contractility. Furthermore, inactivation of PI3Kγ impairs nitric oxide (NO) production by endothelial NO synthase (eNOS), which may lead to increased vascular resistance. Therefore, the aim of our study was to investigate the role of both catalytic and non-catalytic functions of PI3Kγ in the pathogenesis of PH. Methods PI3Kγ knockout mice (PI3Kγ-/-), as well as mice with a catalytically inactive form of PI3Kγ (PI3KγKD/KD) were analysed in vivo using the hypoxia-induced mouse model of PH (21 days at 10% O2 hypoxia (HOX)). To specifically inhibit the non-catalytic function of PI3Kγ, C57BL/6N mice were treated with a mimetic peptide (MP) via intratracheal instillation and studied in the same model. Systolic right ventricular pressure (RVSP) was measured using a Millar pressure catheter inserted via the jugular vein. RV hypertrophy was determined by Fulton's index (RV/LV+S). Results PI3Kγ-/- mice showed significantly increased RVSP after three weeks of hypoxia compared to WT littermate controls (HOX WT 35.64±3.21mmHg vs HOX PI3Kγ-/- 37.04±2.31mmHg;p=0.0049). A significant increase in RVSP was also detected in PI3KγKD/KD and MP-treated mice compared to respective WT controls (HOX WT 34.67±3.83mmHg vs. HOX PI3KγKD/KD 37.9±2.19mmHg;p=0.0404 and HOX vehicle 34.19±2.74mmHg vs. HOX MP 37.61±2.46mmHg;p=0.0111). Under normoxic conditions (NOX), an increased RVSP was already measured in PI3Kγ-/- mice (NOX WT 26.13±1.2mmHg vs. NOX PI3Kγ-/- 28.07±0.88;p=0.0157). Heart rate and systemic blood pressure remained unchanged. A significant increase in RV hypertrophy was only evident for PI3KγKD/KD compared to WT controls (HOX WT 0.38±0.06 vs. HOX PI3KγKD/KD 0.46±0.09mmHg;p=0.0037). Conclusion The results show that both catalytic and non-catalytic inactivation of PI3Kγ in vivo do not counteract the pathogenesis of PH, but conversely enhance it. In this context, reduced phosphorylation of eNOS may play a crucial role, leading to increased vasoconstriction, whereas increased cAMP levels may help the RV to adapt to the increased pressure. The exact mechanisms will be analysed in the future.

Abstract 18479: Lack of Catalytic and Non-Catalytic Function of PI3 Kinase Gamma in Mice Increases Hypoxia-Induced Pulmonary Hypertension

Rationale: Pulmonary hypertension (PH) is characterised by vasoconstriction and remodelling of pulmonary vessels, leading to elevated pulmonary arterial pressure and right ventricular (RV) hypertrophy. PI 3-kinase γ (PI3Kγ) is expressed in leukocytes, cardiomyocytes, and endothelial cells (EC) which are all involved in the pathogenesis of PH. The catalytic function of PI3Kγ is involved in leukocyte recruitment and EC proliferation which are important for vascular remodelling and maladaptive cardiac hypertrophy whereas its non-catalytic function affects cardiac contractility via cAMP signaling. Furthermore, inactivation of PI3Kγ impairs nitric oxide production by eNOS, which may lead to increased vascular resistance. Therefore, we investigated the role of both catalytic and non-catalytic functions of PI3Kγ in the pathogenesis of PH. Methods: PI3Kγ knockout mice (PI3Kγ -/- ), as well as mice with a catalytically inactive form of PI3Kγ (PI3Kγ KD/KD ) were exposed to hypoxia-induced PH (21 days at 10% O 2 hypoxia (HOX)). Systolic right ventricular pressure (RVSP) was measured using a Millar pressure catheter inserted via the jugular vein. RV hypertrophy was determined by Fulton's index. Results: PI3Kγ -/- mice showed significantly increased RVSP after hypoxia compared to WT controls (HOX WT 34.16±3.21 mmHg vs HOX PI3Kγ -/- 37.04±2.31 mmHg; p=0.0049). Already under normoxia (NOX), an increased RVSP was measured in PI3Kγ -/- mice (NOX WT 26.13±1.2mmHg vs. NOX PI3Kγ -/- 28.07±0.88 mmHg, p=0.0157). A significant increase in RVSP was also detected in PI3Kγ KD/KD (HOX WT 34.67±2.02 mmHg vs. HOX PI3Kγ KD/KD 37.95±1.39 mmHg; p=0.0228). Here, a significant increase in RV hypertrophy was evident for PI3Kγ KD/KD mice (HOX WT 0.38±0.06 vs. HOX PI3Kγ KD/KD 0.46±0.09; p=0.0037) which was not detected in PI3Kγ -/- compared to WT controls after hypoxia. Conclusion: The results show that both catalytic and non-catalytic inactivation of PI3Kγ in vivo do not counteract the pathogenesis of PH, but conversely enhance it. In this context, reduced phosphorylation of eNOS may play a crucial role, leading to increased vasoconstriction, whereas increased cAMP levels in PI3Kγ -/- may help the RV to adapt to the increased pressure. The exact mechanisms will be analysed in the future.

Adaptive and maladaptive roles of different angiotensin receptors in the development of cardiac hypertrophy and heart failure.

Angiotensin II (Ang II) is formed by the action of angiotensin-converting enzyme (ACE) in the renin-angiotensin system. This hormone is known to induce cardiac hypertrophy and heart failure and its actions are mediated by the interaction of both pro- and antihypertrophic Ang II receptors (AT1R and AT2R). Ang II is also metabolized by ACE 2 to Ang-(1-7), which elicits the activation of Mas receptors (MasR) for inducing antihypertrophic actions. Since heart failure under different pathophysiological situations is preceded by adaptive and maladaptive cardiac hypertrophy, we have reviewed the existing literature to gain some information regarding the roles of AT1R, AT2R, and MasR in both acute and chronic conditions of cardiac hypertrophy. It appears that the activation of AT1R may be involved in the development of adaptive and maladaptive cardiac hypertrophy as well as subsequent heart failure because both ACE inhibitors and AT1R antagonists exert beneficial effects. On the other hand, the activation of both AT2R and MasR may prevent the occurrence of maladaptive cardiac hypertrophy and delay the progression of heart failure, and thus therapy with different activators of these antihypertrophic receptors under chronic pathological stages may prove beneficial. Accordingly, it is suggested that a great deal of effort should be made to develop appropriate activators of both AT2R and MasR for the treatment of heart failure subjects.

Status of β1-Adrenoceptor Signal Transduction System in Cardiac Hypertrophy and Heart Failure.

Although 1-adrenoceptor ( 1-AR) signal transduction, which maintains cardiac function, is downregulated in failing hearts, the mechanisms for such a defect in heart failure are not fully understood. Since cardiac hypertrophy is invariably associated with heart failure, it is possible that the loss of 1-AR mechanisms in failing heart occurs due to hypertrophic process. In this regard, we have reviewed the information from a rat model of adaptive cardiac hypertrophy and maladaptive hypertrophy at 4 and 24 weeks after inducing pressure overload as well as adaptive cardiac hypertrophy and heart failure at 4 and 24 weeks after inducing volume overload, respectively. Varying degrees of alterations in 1-AR density as well as isoproterenol-induced increases in cardiac function, intracellular -concentration in cardiomyocytes and adenylyl cyclase activity in crude membranes have been reported under these hypertrophic conditions. Adaptive hypertrophy at 4 weeks of pressure or volume overload showed unaltered or augmented increases in the activities of different components of 1-AR signaling. On the other hand, maladaptive hypertrophy due to pressure overload and heart failure due to volume overload at 24 weeks revealed depressions in the activities of 1-AR signal transduction pathway. These observations provide evidence that 1-AR signal system is either unaltered or upregulated in adaptive cardiac hypertrophy and downregulated in maladaptive cardiac hypertrophy or heart failure. Furthermore, the information presented in this article supports the concept that downregulation of 1-AR mechanisms in heart failure or maladaptive cardiac hypertrophy is not due to hypertrophic process per se. It is suggested that a complex mechanism involving the autonomic imbalance may be of a critical importance in determining differential alterations in non-failing and failing hearts.

Real-time influence of intracellular acidification and Na+ /H+ exchanger inhibition on in-cell pyruvate metabolism in the perfused mouse heart: A 31 P-NMR and hyperpolarized 13 C-NMR study.

Disruption of acid-base balance is linked to various diseases and conditions. In the heart, intracellular acidification is associated with heart failure, maladaptive cardiac hypertrophy, and myocardial ischemia. Previously, we have reported that the ratio of the in-cell lactate dehydrogenase (LDH) to pyruvate dehydrogenase (PDH) activities is correlated with cardiac pH. To further characterize the basis for this correlation, these in-cell activities were investigated under induced intracellular acidification without and with Na+ /H+ exchanger (NHE1) inhibition by zoniporide. Male mouse hearts (n= 30) were isolated and perfused retrogradely. Intracellular acidification was performed in two ways: (1) with the NH4 Cl prepulse methodology; and (2) by combining the NH4 Cl prepulse with zoniporide. 31 P NMR spectroscopy was used to determine the intracellular cardiac pH and to quantify the adenosine triphosphate and phosphocreatine content. Hyperpolarized [1-13 C]pyruvate was obtained using dissolution dynamic nuclear polarization. 13 C NMR spectroscopy was used to monitor hyperpolarized [1-13 C]pyruvate metabolism and determine enzyme activities in real time at a temporal resolution of a few seconds using the product-selective saturating excitation approach. The intracellular acidification induced by the NH4 Cl prepulse led to reduced LDH and PDH activities (-16% and -39%, respectively). This finding is in line with previous evidence of reduced myocardial contraction and therefore reduced metabolic activity upon intracellular acidification. Concomitantly, the LDH/PDH activity ratio increased with the reduction in pH, as previously reported. Combining the NH4 Cl prepulse with zoniporide led to a greater reduction in LDH activity (-29%) and to increased PDH activity (+40%). These changes resulted in a surprising decrease in the LDH/PDH ratio, as opposed to previous predictions. Zoniporide alone (without intracellular acidification) did not change these enzyme activities. A possible explanation for the enzymatic changes observed during the combination of the NH4 Cl prepulse and NHE1 inhibition may be related to mitochondrial NHE1 inhibition, which likely negates the mitochondrial matrix acidification. This effect, combined with the increased acidity in the cytosol, would result in an enhanced H+ gradient across the mitochondrial membrane and a temporarily higher pyruvate transport into the mitochondria, thereby increasing the PDH activity at the expense of the cytosolic LDH activity. These findings demonstrate the complexity of in-cell cardiac metabolism and its dependence on intracellular acidification. This study demonstrates the capabilities and limitations of hyperpolarized [1-13 C]pyruvate in the characterization of intracellular acidification as regards cardiac pathologies.

Cardiomyocyte p38 MAPKα suppresses a heart–adipose tissue–neutrophil crosstalk in heart failure development

Although p38 MAP Kinase α (p38 MAPKα) is generally accepted to play a central role in the cardiac stress response, to date its function in maladaptive cardiac hypertrophy is still not unambiguously defined. To induce a pathological type of cardiac hypertrophy we infused angiotensin II (AngII) for 2 days via osmotic mini pumps in control and tamoxifen-inducible, cardiomyocyte (CM)-specific p38 MAPKα KO mice (iCMp38αKO) and assessed cardiac function by echocardiography, complemented by transcriptomic, histological, and immune cell analysis. AngII treatment after inactivation of p38 MAPKα in CM results in left ventricular (LV) dilatation within 48 h (EDV: BL: 83.8 ± 22.5 µl, 48 h AngII: 109.7 ± 14.6 µl) and an ectopic lipid deposition in cardiomyocytes, reflecting a metabolic dysfunction in pressure overload (PO). This was accompanied by a concerted downregulation of transcripts for oxidative phosphorylation, TCA cycle, and fatty acid metabolism. Cardiac inflammation involving neutrophils, macrophages, B- and T-cells was significantly enhanced. Inhibition of adipose tissue lipolysis by the small molecule inhibitor of adipocytetriglyceride lipase (ATGL) Atglistatin reduced cardiac lipid accumulation by 70% and neutrophil infiltration by 30% and went along with an improved cardiac function. Direct targeting of neutrophils by means of anti Ly6G-antibody administration in vivo led to a reduced LV dilation in iCMp38αKO mice and an improved systolic function (EF: 39.27 ± 14%). Thus, adipose tissue lipolysis and CM lipid accumulation augmented cardiac inflammation in iCMp38αKO mice. Neutrophils, in particular, triggered the rapid left ventricular dilatation. We provide the first evidence that p38 MAPKα acts as an essential switch in cardiac adaptation to PO by mitigating metabolic dysfunction and inflammation. Moreover, we identified a heart–adipose tissue–immune cell crosstalk, which might serve as new therapeutic target in cardiac pathologies.

Inactivation of catalytic and non-catalytic PI3 kinase gamma functions enhances hypoxia-induced pulmonary hypertension in mice

Abstract Rationale Pulmonary hypertension (PH) is a pulmonary vascular disease characterised by chronically elevated pulmonary arterial mean pressure, increased pulmonary vascular resistance and right ventricular (RV) dysfunction and hypertrophy. The pathogenesis includes vasoconstriction of pulmonary vessels and pulmonary vascular remodelling. Phosphatidylinositol 3'-kinase γ (PI3Kγ) is highly expressed in leukocytes, endothelial cells (EC) and cardiomyocytes, which are involved in the pathogenesis of PH. The catalytic function of PI3Kγ has been shown to be associated with numerous processes that are potentially important for both vascular remodelling and maladaptive cardiac hypertrophy, including leukocyte recruitment and endothelial cell proliferation and survival. However, inhibition of non-catalytic PI3Kγ function results in reduced nitric oxide (NO) production by endothelial NO synthase (eNOS), which may lead to increased vascular resistance. Therefore, the aim of our study was to investigate the role of both catalytic and non-catalytic PI3Kγ functions in the pathogenesis of PH. Methods The importance of PI3Kγ in the pathogenesis of PH was analysed in vivo using the hypoxia-induced mouse model of PH (21 days at 10% O2 hypoxia (HOX)). PI3Kγ knockout mice (PI3Kγ−/−), as well as mice expressing a catalytically inactive form of PI3Kγ (PI3KγKD/KD) were investigated. To specifically inhibit the non-catalytic function of PI3Kγ, C57BL/6N mice were treated with a blocking peptide via intratracheal instillation and studied in the same model. Subsequently, systolic right ventricular pressure (RVSP) was measured using a Millar pressure catheter inserted via the jugular vein. In vitro, western blotting was used to investigate the phosphorylation of eNOS (Ser1177) in pulmonary ECs from wild type (WT), PI3Kγ−/− and PI3KγKD/KD mice. Results PI3Kγ−/− mice showed significantly increased RVSP after three weeks of hypoxia compared with WT controls (HOX WT 34.16±3.47mmHg vs. HOX PI3Kγ−/− 37.04±2.43mmHg; p=0.005; n≥7). A significant increase in RVSP was also detected in PI3KγKD/KD and blocking peptide-treated mice (HOX WT 34.67±2.02mmHg vs. HOX PI3KγKD/KD 37.95±1.39mmHg; p=0.023; n≥5 and HOX vehicle 34.19±2.74mmHg vs. HOX blocking peptide 37.61±2.46mmHg; p=0.011; n≥9). Heart rate, as well as systemic blood pressure, remained unchanged. Under normoxic conditions, no difference in RVSP between the groups could be measured. Interestingly, western blot detected reduced phosphorylation of eNOS after stimulation with fetal calf serum in PI3Kγ−/− ECs compared to WT and PI3KγKD/KD. Conclusion These results show that blunting of both catalytic and non-catalytic functions of PI3Kγ in vivo do not prevent the pathogenesis of PH, but conversely enhance hypoxia-induced elevation of RVSP. In this context, reduced phosphorylation of eNOS may play a crucial role, leading to increased vasoconstriction. Additional studies are required to evaluate the underlying mechanisms in detail. Funding Acknowledgement Type of funding sources: Foundation. Main funding source(s): DFG Deutsche Forschungsgemeinschaft

Statins Protect Against Early Stages of Doxorubicin-induced Cardiotoxicity Through the Regulation of Akt Signaling and SERCA2

BackgroundDoxorubicin-induced cardiomyopathy (DICM) is one of the complications that can limit treatment for a significant number of cancer patients. In animal models, the administration of statins can prevent the development of DICM. Therefore, the use of statins with anthracyclines potentially could enable cancer patients to complete their chemotherapy without added cardiotoxicity. The precise mechanism mediating the cardioprotection is not well understood. The purpose of this study is to determine the molecular mechanism by which rosuvastatin confers cardioprotection in a mouse model of DICM. MethodsRosuvastatin was intraperitoneally administered into adult male mice at 100 μg/kg daily for 7 days, followed by a single intraperitoneal doxorubicin injection at 10 mg/kg. Animals continued to receive rosuvastatin daily for an additional 14 days. Cardiac function was assessed by echocardiography. Optical calcium mapping was performed on retrograde Langendorff perfused isolated hearts. Ventricular tissue samples were analyzed by immunofluorescence microscopy, Western blotting, and quantitative polymerase chain reaction. ResultsExposure to doxorubicin resulted in significantly reduced fractional shortening (27.4% ± 1.11% vs 40% ± 5.8% in controls; P < 0.001) and re-expression of the fetal gene program. However, we found no evidence of maladaptive cardiac hypertrophy or adverse ventricular remodeling in mice exposed to this dose of doxorubicin. In contrast, rosuvastatin-doxorubicin-treated mice maintained their cardiac function (39% ± 1.26%; P < 0.001). Mechanistically, the effect of rosuvastatin was associated with activation of Akt and phosphorylation of phospholamban with preserved sarcoplasmic/endoplasmic reticulum Ca2+ transporting 2 (SERCA2)-mediated Ca2+ reuptake. These effects occurred independently of perturbations in ryanodine receptor 2 function. ConclusionsRosuvastatin counteracts the cardiotoxic effects of doxorubicin by directly targeting sarcoplasmic calcium cycling.

The focal adhesion protein β-parvin controls cardiomyocyte shape and sarcomere assembly in response to mechanical load

Physiological and pathological cardiac stress induced by exercise and hypertension, respectively, increase the hemodynamic load for the heart and trigger specific hypertrophic signals in cardiomyocytes leading to adaptive or maladaptive cardiac hypertrophy responses involving a mechanosensitive remodeling of the contractile cytoskeleton. Integrins sense load and have been implicated in cardiac hypertrophy, but how they discriminate between the two types of cardiac stress and translate mechanical loads into specific cytoskeletal signaling pathways is not clear. Here, we report that the focal adhesion protein β-parvin is highly expressed in cardiomyocytes and facilitates the formation of cell protrusions, the serial assembly of newly synthesized sarcomeres, and the hypertrophic growth of neonatal rat ventricular cardiomyocytes (NRVCs) invitro. In addition, physiological mechanical loading of NRVCs by either the application of cyclic, uni-axial stretch, or culture on physiologically stiff substrates promotes NRVC elongation in a β-parvin-dependent manner, which is achieved by binding of β-parvin to α/β-PIX, which in turn activates Rac1. Importantly, loss-of-function studies in mice also revealed that β-parvin is essential for the exercise-induced cardiac hypertrophy response invivo. Our results identify β-parvin as a novel mechano-responsive signaling hub in hypertrophic cardiomyocytes that drives cell elongation in response to physiological mechanical loads.

Decreased Native T1 Values and Impaired Myocardial Contractility in Anabolic Steroid Users.

Anabolic androgenic steroid (AAS) abuse leads to myocardial toxicity. Human studies are conflicting about the myocardial fibrosis in AAS users. We evaluated cardiac tissue characterization, left ventricle (LV) function, and cardiac structure by cardiovascular magnetic resonance (CMR). Twenty strength-trained AAS users (AASU) aged 29±5 yr, 20 strength-trained AAS nonusers (AASNU), and 7 sedentary controls (SC) were enrolled. Native T1 mapping, late-gadolinium enhancement (LGE), extracellular volume (ECV), and myocardial strain were evaluated. AASU showed lower Native T1 values than AASNU (888±162 vs. 1020±179 ms p=0.047). Focal myocardial fibrosis was found in 2 AASU. AASU showed lower LV radial strain (30±8 vs. 38±6%, p<0.01), LV circumferential strain (-17±3 vs. -20±2%, p<0.01), and LV global longitudinal strain (-17±3 vs. -20±3%, p<0.01) than AASNU by CMR. By echocardiography, AASU demonstrated lower 4-chamber longitudinal strain than AASNU (-15±g3 vs. -18±2%, p=0.03). ECV was similar among AASU, AASNU, and SC (28±10 vs. 28±7 vs. 30±7%, p=0.93). AASU had higher LV mass index than AASNU and SC (85±14 vs. 64±8 vs. 58±5 g/m2, respectively, p<0.01). AAS abuse may be linked to decreased myocardial native T1 values, impaired myocardial contractility, and focal fibrosis. These alterations may be associated with maladaptive cardiac hypertrophy in young AAS users.

Protective Effects of Thyroid Hormone Deprivation on Progression of Maladaptive Cardiac Hypertrophy and Heart Failure.

Purpose: Thyroid hormones (TH) play a central role for cardiac function. TH influence heart rate and cardiac contractility, and altered thyroid function is associated with increased cardiovascular morbidity and mortality. The precise role of TH in onset and progression of heart failure still requires clarification.Methods: Chronic left ventricular pressure overload was induced in mouse hearts by transverse aortic constriction (TAC). One week after TAC, alteration of TH status was induced and the impact on cardiac disease progression was studied longitudinally over 4 weeks in mice with hypo- or hyperthyroidism and was compared to euthyroid TAC controls. Serial assessment was performed for heart function (2D M-mode echocardiography), heart morphology (weight, fibrosis, and cardiomyocyte cross-sectional area), and molecular changes in heart tissues (TH target gene expression, apoptosis, and mTOR activation) at 2 and 4 weeks.Results: In diseased heart, subsequent TH restriction stopped progression of maladaptive cardiac hypertrophy and improved cardiac function. In contrast and compared to euthyroid TAC controls, increased TH availability after TAC propelled maladaptive cardiac growth and development of heart failure. This was accompanied by a rise in cardiomyocyte apoptosis and mTOR pathway activation.Conclusion: This study shows, for the first time, a protective effect of TH deprivation against progression of pathological cardiac hypertrophy and development of congestive heart failure in mice with left ventricular pressure overload. Whether this also applies to the human situation needs to be determined in clinical studies and would infer a critical re-thinking of management of TH status in patients with hypertensive heart disease.

A peptide of the N terminus of GRK5 attenuates pressure-overload hypertrophy and heart failure.

Aberrant changes in gene expression underlie the pathogenesis and progression of pressure-overload heart failure, leading to maladaptive cardiac hypertrophy, ventricular remodeling, and contractile dysfunction. Signaling through the G protein Gq triggers maladaptation and heart failure, in part through the activation of G protein-coupled receptor kinase 5 (GRK5). Hypertrophic stimuli induce the accumulation of GRK5 in the nuclei of cardiomyocytes, where it regulates pathological gene expression through multiple transcription factors including NFAT. The nuclear targeting of GRK5 is mediated by an amino-terminal (NT) domain that binds to calmodulin (CaM). Here, we sought to prevent GRK5-mediated pathology in pressure-overload maladaptation and heart failure by expressing in cardiomyocytes a peptide encoding the GRK5 NT (GRK5nt) that encompasses the CaM binding domain. In cultured cardiomyocytes, GRK5nt expression abrogated Gq-coupled receptor-mediated hypertrophy, including attenuation of pathological gene expression and the transcriptional activity of NFAT and NF-κB. We confirmed that GRK5nt bound to and blocked Ca2+-CaM from associating with endogenous GRK5, thereby preventing GRK5 nuclear accumulation after pressure overload. We generated mice that expressed GRKnt in a cardiac-specific fashion (TgGRK5nt mice), which exhibited reduced cardiac hypertrophy, ventricular dysfunction, pulmonary congestion, and cardiac fibrosis after chronic transverse aortic constriction. Together, our data support a role for GRK5nt as an inhibitor of pathological GRK5 signaling that prevents heart failure.

Unraveling the Role of 12- and 20- HETE in Cardiac Pathophysiology: G-Protein-Coupled Receptors, Pharmacological Inhibitors, and Transgenic Approaches.

Arachidonic acid-derived lipid mediators play crucial roles in the development and progression of cardiovascular diseases. Eicosanoid metabolites generated by lipoxygenases and cytochrome P450 enzymes produce several classes of molecules, including the epoxyeicosatrienoic acid (EET) and hydroxyeicosatetraenoic acids (HETE) family of bioactive lipids. In general, the cardioprotective effects of EETs have been documented across a number of cardiac diseases. In contrast, members of the HETE family have been shown to contribute to the pathogenesis of ischemic cardiac disease, maladaptive cardiac hypertrophy, and heart failure. The net effect of 12(S)- and 20-HETE depends upon the relative amounts generated, ratio of HETEs:EETs produced, timing of synthesis, as well as cellular and subcellular mechanisms activated by each respective metabolite. HETEs are synthesized by and affect multiple cell types within the myocardium. Moreover, cytochrome P450-derived and lipoxygenase- derived metabolites have been shown to directly influence cardiac myocyte growth and the regulation of cardiac fibroblasts. The mechanistic data uncovered thus far have employed the use of enzyme inhibitors, HETE antagonists, and the genetic manipulation of lipid-producing enzymes and their respective receptors, all of which influence a complex network of outcomes that complicate data interpretation. This review will summarize and integrate recent findings on the role of 12(S)-/20-HETE in cardiac diseases.

G protein-coupled receptor kinase 5 (GRK5) contributes to impaired cardiac function and immune cell recruitment in post-ischemic heart failure.

Aims Myocardial infarction (MI) is the most common cause of heart failure (HF) worldwide. G protein-coupled receptor kinase 5 (GRK5) is upregulated in failing human myocardium and promotes maladaptive cardiac hypertrophy in animal models. However, the role of GRK5 in ischemic heart disease is still unknown. In this study, we evaluated whether myocardial GRK5 plays a critical role post-MI in mice and included the examination of specific cardiac immune and inflammatory responses.Methods and results Cardiomyocyte-specific GRK5 overexpressing transgenic mice (TgGRK5) and non-transgenic littermate control (NLC) mice as well as cardiomyocyte-specific GRK5 knockout mice (GRK5cKO) and wild type (WT) were subjected to MI and, functional as well as structural changes together with outcomes were studied. TgGRK5 post-MI mice showed decreased cardiac function, augmented left ventricular dimension and decreased survival rate compared to NLC post-MI mice. Cardiac hypertrophy and fibrosis as well as fetal gene expression were increased post-MI in TgGRK5 compared to NLC mice. In TgGRK5 mice, GRK5 elevation produced immuno-regulators that contributed to the elevated and long-lasting leukocyte recruitment into the injured heart and ultimately to chronic cardiac inflammation. We found an increased presence of pro-inflammatory neutrophils and macrophages as well as neutrophils, macrophages and T-lymphocytes at 4-days and 8-weeks respectively post-MI in TgGRK5 hearts. Conversely, GRK5cKO mice were protected from ischemic injury and showed reduced early immune cell recruitment (predominantly monocytes) to the heart, improved contractility and reduced mortality compared to WT post-MI mice. Interestingly, cardiomyocyte-specific GRK2 transgenic mice did not share the same phenotype of TgGRK5 mice and did not have increased cardiac leukocyte migration and cytokine or chemokine production post-MI.Conclusions Our study shows that myocyte GRK5 has a crucial and GRK-selective role on the regulation of leucocyte infiltration into the heart, cardiac function and survival in a murine model of post-ischemic HF, supporting GRK5 inhibition as a therapeutic target for HF.