Cardiac Effects of Attenuating Gsα - Dependent Signaling.

AIMS: Inhibition of ß-adrenergic signalling plays a key role in treatment of heart failure. Gsα is essential for ß-adrenergic signal transduction. In order to reduce side-effects of beta-adrenergic inhibition diminishing ß-adrenergic signalling in the heart at the level of Gsα is a promising option. METHODS AND RESULTS: We analyzed the influence of Gsα on regulation of myocardial function and development of cardiac hypertrophy, using a transgenic mouse model (C57BL6/J mice) overexpressing a dominant negative Gsα-mutant under control of the α-MHC-promotor. Cardiac phenotype was characterized in vivo and in vitro and under acute and chronic ß-adrenergic stimulation. At rest, Gsα-DN-mice showed bradycardia (602 ± 13 vs. 660 ± 17 bpm, p<0.05) and decreased dp/dtmax (5037 ± 546- vs. 6835 ± 505 mmHg/s, p = 0.02). No significant differences were found regarding ejection fraction, heart weight and cardiomyocyte size. ß-blockade by propranolol revealed no baseline differences of hemodynamic parameters between wildtype and Gsα-DN-mice. Acute adrenergic stimulation resulted in decreased ß-adrenergic responsiveness in Gsα-DN-mice. Under chronic adrenergic stimulation, wildtype mice developed myocardial hypertrophy associated with increase of LV/BW-ratio by 23% (4.4 ± 0.2 vs. 3.5 ± 0.1 mg/g, p<0.01) and cardiac myocyte size by 24% (14927 ± 442 px vs. 12013 ± 583 px, p<0.001). In contrast, both parameters were unchanged in Gsα-DN-mice after chronic isoproterenol stimulation. CONCLUSION: Overexpression of a dominant negative mutant of Gsα leads to decreased ß-adrenergic responsiveness and is protective against isoproterenol-induced hypertrophy. Thus, Gsα-DN-mice provide novel insights into ß-adrenergic signal transduction and its modulation in myocardial overload and failure.

DYRK2 negatively regulates cardiomyocyte growth by mediating repressor function of GSK-3ß on eIF2Bε.

BACKGROUND: A prerequisite of hypertrophic response of the myocardium is an increase in protein synthesis. A central regulator of translation initiation is Eukaryotic initiation factor 2B (eIF2B). Here we assessed the hypothesis that regulation of protein synthesis via eIF2Bε is essential to cardiac hypertrophic response in vivo. METHODS: Two transgenic mouse lines were generated with cardiac restricted overexpression of eIF2Bε or its mutant eIF2Bε-eIFS(535)A, which cannot be inactivated by phosphorylation through GSK-3ß. RESULTS: (1) Under baseline conditions eIF2Bε transgenic mice showed no difference in cardiac phenotype compared to wild type, whereas in the mutant eIF2Bε-S(535)A an increase in LV/tibia length (7.5 ± 0.4 mg/mm vs. 6.2 ± 0.2 mg/mm, p<0.001) and cardiomyocyte cross sectional area (13004 ± 570 vs. 10843 ± 347 RU, p<0.01) was observed. (2) Cardiac overexpression of eIF2Bε did not change the response of the heart to pathologic stress induced by chronic isoproterenol treatment. (3) Cardiac overexpression of the eIF2Bε transgene was followed by overexpression of DYRK2 which is known to prime the inhibitory action of GSK-3ß on eIF2Bε, while DYRK1A and GSK-3ß itself were not increased. (4) In C57BL/6 mice after 48 h of isoproterenol-stimulation or aortic banding, eIF2Bε was increased and DYRK2 was concomitantly decreased. (5) In line with these in vivo findings, siRNA knockdown of DYRK2 in cultured cardiomyocytes resulted in decreased levels of p(S535)- eIF2Bε, (6) whereas adenoviral induced overexpression of DYRK2 was accompanied by clearly increased phosphorylation of eIF2Bε, indicating a coordinated response pattern (7) Adenoviral induced overexpression of DYRK2 leads to significantly reduced cardiomyocyte size and diminishes hypertrophic response to adrenergic stimulation. CONCLUSIONS: The interaction of GSK-3ß and its priming kinase DYRK2 regulate the activity of eIF2Bε in cardiac myocytes. DYRK2 is a novel negative regulator of cardiomyocyte growth. DYRK2 could serve as a therapeutic option to regulate myocardial growth.

High-sensitive Troponin T increase after exercise in patients with pulmonary arterial hypertension.

BACKGROUND: The current study aimed to investigate the release of myocardial high-sensitive Troponin T (hsTnT) in patients with pulmonary arterial hypertension (PAH) in response to maximal physical exercise. METHODS: In 24 patients with PAH, symptom-limited cardiopulmonary exercise testing was performed. hsTnT was measured by the novel hsTnT assay with a lower limit of detection of 2 ng/L and a total imprecision of less than 10% at the 99th percentile value. hsTnT was related to NT-proBNP, WHO functional class and right ventricular (RV) function. Serial measurement was performed before and 30 min, 180 min, and 300 min after exercise. Healthy volunteers served as a control group. RESULTS: In 21 PAH patients, hsTnT levels were detectable before exercise with a close correlation between hsTnT and NT-proBNP. hsTnT was detectable in all PAH patients after exercise and significantly increased from 7.5 ng/L at baseline to 14.62 ng/L after 300 min, whereas levels of NT-proBNP remained constant with time. CONCLUSIONS: Using the novel hsTnT assay, the current study provides first evidence that hsTnT levels increase in PAH patients after maximal physical exercise, while levels of other biomarkers remain constant after exercise testing. This might provide new insights into pathophysiology and individual risk assessment in patients with PAH.

Wnt signaling is critical for maladaptive cardiac hypertrophy and accelerates myocardial remodeling.

The evolutionary conserved Wnt signaling pathway regulates cardiogenesis. However, members of the Wnt pathway are also expressed in the adult heart. Although Wnt-signaling is quiescent under normal conditions, we noticed activation on pathological stress of the heart, such as chronic afterload increase. To examine the role of Wnt signaling on the postnatal heart, we modified the expression and function of the Wnt regulator dishevelled 1 (Dvl-1) both in transgenic mice with cardiac-specific overexpression of Dvl-1 (Dvl-1-Tg) and in cultured cardiac myocytes. Dvl-1-Tg mice (3 months) had severe cardiac hypertrophy (heart weight:body weight ratio: 5.2+/-0.3 mg/g wild-type [WT] versus 6.4+/-0.7 mg/g Dvl-1-Tg; P<0.01), an increase in cardiomyocyte size (86% increase in Dvl-1-Tg compared with WT; P<0.01) and marked raise of atrial natriuretic factor expression (12-fold increase versus WT; P<0.01). Hypertrophy was associated with left ventricular dilatation in Dvl-1-Tg and a reduction of ejection fraction (4.4+/-0.1 mm versus 5.5+/-0.2 mm, 80+/-2% and 43+/-4% in WT versus Dvl-1-Tg, respectively; P<0.01). Transgenic animals died prematurely before 6 months of age. Both canonical as well as noncanonical Wnt signaling branches were activated in the Dvl-1-Tg animals. Small interfering RNA-mediated depletion of Dvl-1 was used to further characterize the role of Dvl-1 in cardiac myocytes. Whereas baseline parameters were unaltered, beta-adrenergic hypertrophic response was abrogated in Dvl-1 knockdown cardiac myocytes, indicating a mandatory role in beta-adrenergic stimulation. Therefore, activation of Wnt signaling is sufficient and critical for the induction of myocardial hypertrophy and cardiomyopathy.

Activation of PPARgamma by pioglitazone does not attenuate left ventricular hypertrophy following aortic banding in rats.

Sustained left ventricular hypertrophy (LVH) accelerates cardiac dysfunction and heart failure. Previous reports have suggested that activation of the peroxisome proliferator-activated receptor gamma (PPARgamma)-dependent pathway is involved in development of cardiac hypertrophy. Thiazolidinediones (TZDs) such as pioglitazone activate PPARgamma and are clinically used as antidiabetics. Given inconsistent reports regarding effects of TZDs on LVH, we examined in the present study the influence of pioglitazone on LVH in a rat model of aortic banding. Aortic banding was induced in rats by clipping the ascending aorta. Animals received pioglitazone (3 mg/kg/day) or placebo. Echocardiographic, hemodynamic, histological, and biochemical measurements were performed after 2 and 4 weeks. Pressure gradient was comparable between pioglitazone- and placebo-treated animals after 2 and 4 weeks. Left ventricular function was not different between the groups. In sham as well as in banded animals, LV/body weight ratio was increased in pioglitazone- as compared to placebo-treated animals after 2 and 4 weeks. Furthermore, an increase in myocyte size and atrial natriuretic factor was observed in pioglitazone- compared to placebo-treated animals 4 weeks after aortic banding as well. The results of this study demonstrate that activation of PPARgamma via pioglitazone does not protect the myocardium from pressure overload-induced LVH in a rat model of aortic banding. The findings rather indicate a pro-hypertrophic effect of pioglitazone treatment after aortic banding.

Myocardial left ventricular dysfunction in patients with systemic lupus erythematosus: new insights from tissue Doppler and strain imaging.

OBJECTIVE: Systemic lupus erythematosus (SLE) is associated with high cardiovascular morbidity and mortality. Cardiovascular involvement is frequently underestimated by routine imaging techniques. Our aim was to determine if new echocardiographic imaging modalities like tissue Doppler (TDI), strain rate (SRR), and strain (SRI) imaging detect abnormalities in left ventricular (LV) function in asymptomatic patients with SLE. METHODS: Sixty-seven young patients with SLE (mean age 42 +/- 10 yrs) without typical symptoms or signs of heart failure or angina, and a matched healthy control group (n = 40), underwent standard transthoracic echocardiography, TDI, SRR, and SRI imaging of the LV as well as assessment of disease characteristics. RESULTS: Despite findings within the normal range on routine standard 2-dimensional echocardiography, SLE was associated with significantly impaired systolic and diastolic myocardial velocities of the LV measured by TDI [mean global TDI: systolic (s): 2.9 +/- 0.9 vs 3.9 +/- 0.7 cm/s, p < 0.05; early (e): 4.3 +/- 1.5 vs 6.3 +/- 1.3 cm/s, p < 0.05; late (a): 2.9 +/- 0.8 vs 3.4 +/- 0.8 cm/s, p < 0.05; values +/- SD); SRR (s: -0.8 +/- 0.1 vs -1.1 +/- 0.1 s(-1); e: 1.1 +/- 0.2 vs 1.6 +/- 0.3 s(-1); a: 0.7 +/- 0.1 vs 1.0 +/- 0.2 s(-1); all p < 0.05); and SR (-15.11 +/- 2.2% vs -19.7 +/- 1.9%; p < 0.05) compared to the control group. Further, elevated disease activity, measured with the ECLAM and the SLEDAI score, resulted in significantly lower values for LV longitudinal function measured by SRR and SR, but not by TDI. CONCLUSION: SLE is associated with a significant impairment of systolic and diastolic LV longitudinal function in patients without cardiac symptoms. New imaging modalities provide earlier insight into cardiovascular involvement in SLE and seem to be superior to standard echocardiography to detect subclinical myocardial disease.

Depletion of mammalian target of rapamycin (mTOR) via siRNA mediated knockdown leads to stabilization of beta-catenin and elicits distinct features of cardiomyocyte hypertrophy.

Cardiac myocyte growth is under differential control of mammalian target of rapamycin (mTOR) and glycogen-synthase-kinase-3beta (GSK3beta). Whereas active GSK3beta negatively regulates growth and down-regulates cellular protein synthesis, activation of the mTOR pathway promotes protein expression and cell growth. Here we report that depletion of mTOR via siRNA mediated knockdown causes marked down-regulation of GSK3beta protein in cardiac myocytes. As a result, GSK3beta target protein beta-catenin becomes stabilized and translocates into the nucleus. Moreover, mTOR knockdown leads to increase in cardiac myocyte surface area and produces an up-regulation of the fetal gene program. Our findings suggest a new type of convergence of mTOR and GSK3beta activities, indicating that GSK3beta-dependent stabilization of beta-catenin in cardiac myocytes is influenced by mTOR.

Beneficial effects of Mammalian target of rapamycin inhibition on left ventricular remodeling after myocardial infarction.

OBJECTIVES: The extent of adverse myocardial remodeling contributes essentially to the prognosis after myocardial infarction (MI). In this study we investigated whether inhibition of "mammalian target of rapamycin" (mTOR) attenuates left ventricular (LV) remodeling after MI. BACKGROUND: Therapeutic strategies to inhibit remodeling are currently limited to inhibition of neurohumoral activation. The mTOR-dependent signaling mechanisms are centrally involved in remodeling processes and provide new therapeutic opportunities. METHODS: Everolimus (RAD) treatment was initiated on the day after or 3 days after induction of myocardial infarction (MI) in rats. RESULTS: After 28 days, RAD-treated animals had reduced post-MI remodeling, with improved LV function and smaller LV end-diastolic diameters (8.9 + or - 0.3 mm vs. 11.4 + or - 0.2 mm, p < 0.05), end-diastolic volumes (304 + or - 30 microl vs. 414 + or - 16 microl, p < 0.05), and cardiac myocyte size (-40% vs. vehicle, p < 0.05). Infarct size was significantly reduced compared with vehicle-treated animals. The mTOR inhibition increased autophagy and concomitantly decreased proteasome activity in the border zone of the infarcted myocardium. Measurement of autophagic flux demonstrated that RAD did not decrease autophagosome clearance. When RAD treatment was initiated 3 days after MI, adverse remodeling was still attenuated and increased autophagy was still present. Sustained improvement of LV function was observed 3 months after MI, even when RAD treatment was discontinued after 1 month. CONCLUSIONS: Inhibition of mTOR is a potential therapeutic strategy to limit infarct size and to attenuate adverse LV remodeling after MI.