Georg Ertl

MicroRNAs in the human heart : A clue to fetal gene reprogramming in heart failure

Background— Chronic heart failure is characterized by left ventricular remodeling and reactivation of a fetal gene program; the underlying mechanisms are only partly understood. Here we provide evidence that cardiac microRNAs, recently discovered key regulators of gene expression, contribute to the transcriptional changes observed in heart failure. Methods and Results— Cardiac transcriptome analyses revealed striking similarities between fetal and failing human heart tissue. Using microRNA arrays, we discovered profound alterations of microRNA expression in failing hearts. These changes closely mimicked the microRNA expression pattern observed in fetal cardiac tissue. Bioinformatic analysis demonstrated a striking concordance between regulated messenger RNA expression in heart failure and the presence of microRNA binding sites in the respective 3 untranslated regions. Messenger RNAs upregulated in the failing heart contained preferentially binding sites for downregulated microRNAs and vice versa. Mechanistically, transfection of cardiomyocytes with a set of fetal microRNAs induced cellular hypertrophy as well as changes in gene expression comparable to the failing heart. Conclusions— Our data support a novel mode of regulation for the transcriptional changes in cardiac failure. Reactivation of a fetal microRNA program substantially contributes to alterations of gene expression in the failing human heart.

Progressive left ventricular dysfunction and remodeling after myocardial infarction. Potential mechanisms and early predictors.

BackgroundLeft ventricular enlargement and the development of chronic heart failure are potent predictors of survival in patients after myocardial infarction. Prospective studies relating progressive ventricular enlargement in individual patients to global and regional cardiac dysfunction and the onset of late chronic heart failure are not available. It was the aim of this study to define the relation between left ventricular dilatation and global and regional cardiac dysfunction and to identify early predictors of enlargement and chronic heart failure in patients after myocardial infarction. Methods and ResultsLeft ventricular volumes, regional area shrinkage fraction in 18 predefined sectors (gated single photon emission computed tomography), global ejection fraction, and hemodynamics at rest and during exercise (supine bicycle, 50 W, 4 minutes, Swan-Ganz catheter) were assessed prospectively 4 days, 4 weeks, 6 months, and 1.5 and 3 years after first myocardial infarction. Seventy patients were assigned to groups with progressive, limited, or no dilatation. Patients without dilatation (n=38) maintained normal volumes and hemodynamics until 3 years. With limited dilatation (n= 18), left ventricular volume increased up to 4 weeks after infarction and stabilized thereafter, depressed stroke volume was restored 4 weeks after infarction and then remained stable at rest. Wedge pressure during exercise, however, progressively increased. With progressive dilatation (n= 14), depressed cardiac and stroke indexes were also restored by 4 weeks but progressively deteriorated thereafter. Area shrinkage fraction as an estimate of regional left ventricular function in normokinetic sectors at 4 days gradually deteriorated during 3 years, but hypokinetic and dyskinetic sectors remained unchanged. Global ejection fraction fell after 1.5 years, whereas right atrial pressure, wedge pressure, and systemic vascular resistance increased. By multivariate analysis, ejection fraction and stroke index at 4 days, ventriculographic infarct size, infarct location, and Thrombolysis in Myocardial Infarction trial grade of infarct artery perfusion were significant predictors of progressive ventricular enlargement and chronic dysfunction. ConclusionAlmost 26% of patients may develop limited leftventricular dilatation within 4 weeks after first infarction, which helps to restore cardiac index and stroke index at rest and to preserve exercise performance and therefore remains compensatory. A somewhat smaller group (20%) develops progressive structural leftventricular dilatation, which is compensatory at first, then progresses to noncompensatory dilatation, and finally results in severe global left ventricular dysfunction. In these patients, depression of global ejection fraction probably results from impairment of functionof initially normally contracting myocardium. Early predictors from multivariate analysis allow identification of patients at high risk for progressive left ventricular dilatation and chronic ventricular dysfunction within 4 weeks after acute infarction.

Myocardial Phosphocreatine-to-ATP Ratio Is a Predictor of Mortality in Patients With Dilated Cardiomyopathy

BACKGROUND In patients with heart failure due to dilated cardiomyopathy, cardiac energy metabolism is impaired, as indicated by a reduction of the myocardial phosphocreatine-to-ATP ratio, measured noninvasively by 31P-MR spectroscopy. The purpose of this study was to test whether the phosphocreatine-to-ATP ratio also offers prognostic information in terms of mortality prediction as well as how this index compares with well-known mortality predictors such as left ventricular ejection fraction (LVEF) or New York Heart Association (NYHA) class. METHODS AND RESULTS Thirty-nine patients with dilated cardiomyopathy were followed up for 928+/-85 days (2.5 years). At study entry, LVEF and NYHA class were determined, and the cardiac phosphocreatine-to-ATP ratio was measured by localized 31P-MR spectroscopy of the anterior myocardium. During the study period, total mortality was 26%. Patients were divided into two groups, one with a normal phosphocreatine-to-ATP ratio (>1.60; mean+/-SE, 1.98+/-0.07; n=19; healthy volunteers: 1.94+/-0.11, n=30) and one with a reduced phosphocreatine-to-ATP ratio (<1.60; 1.30+/-0.05; n=20). At re-evaluation (mean, 2.5 years), 8 of 20 patients with reduced phosphocreatine-to-ATP ratios had died, all of cardiovascular causes (total and cardiovascular mortality, 40%). Of the 19 patients with normal phosphocreatine-to-ATP ratios, 2 had died (total mortality, 11%), one of cardiovascular causes (cardiovascular mortality, 5%). Kaplan-Meier analysis showed significantly reduced total (P=.036) and cardiovascular (P=.016) mortality for patients with normal versus patients with low phosphocreatine-to-ATP ratios. A Cox model for multivariate analysis showed that the phosphocreatine-to-ATP ratio and NYHA class offered significant independent prognostic information on cardiovascular mortality. CONCLUSIONS The myocardial phosphocreatine-to-ATP ratio, measured noninvasively with 31P-MR spectroscopy, is a predictor of both total and cardiovascular mortality in patients with dilated cardiomyopathy.

Impact of Myocardial Fibrosis in Patients With Symptomatic Severe Aortic Stenosis

Background— In this prospective follow-up study, the effect of myocardial fibrosis on myocardial performance in symptomatic severe aortic stenosis was investigated, and the impact of fibrosis on clinical outcome after aortic valve replacement (AVR) was estimated. Methods and Results— Fifty-eight consecutive patients with isolated symptomatic severe aortic stenosis underwent extensive baseline characterization before AVR. Standard and tissue Doppler echocardiography and cardiac magnetic resonance imaging (late-enhancement imaging for replacement fibrosis) were performed at baseline and 9 months after AVR. Endomyocardial biopsies were obtained intraoperatively to determine the degree of myocardial fibrosis. Patients were analyzed according to the severity of interstitial fibrosis in cardiac biopsies (severe, n=21; mild, n=15; none, n=22). The extent of histologically determined cardiac fibrosis at baseline correlated closely with New York Heart Association functional class and markers of longitudinal systolic function (all P<0.001) but not global ejection fraction or aortic valve area. Nine months after AVR, the degree of late enhancement remained unchanged, implying that AVR failed to reduce the degree of replacement fibrosis. Patients with no fibrosis experienced a marked improvement in New York Heart Association class from 2.8±0.4 to 1.4±0.5 (P<0.001). Only parameters of longitudinal systolic function predicted this functional improvement. Four patients with severe fibrosis died during follow-up, but no patient from the other groups died. Conclusions— Myocardial fibrosis is an important morphological substrate of postoperative clinical outcome in patients with severe aortic stenosis and was not reversible after AVR over the 9 months of follow-up examined in this study. Because markers of longitudinal systolic function appear to indicate sensitively both the severity of myocardial fibrosis and the clinical outcome, they may prove valuable for preoperative risk assessment in patients with aortic stenosis.

Improvement of Cardiac Function During Enzyme Replacement Therapy in Patients With Fabry Disease A Prospective Strain Rate Imaging Study

Background—Enzyme replacement therapy (ERT) has been shown to enhance microvascular endothelial globotriaosylceramide clearance in the hearts of patients with Fabry disease. Whether these results can be translated into an improvement of myocardial function has yet to be demonstrated. Methods and Results—Sixteen patients with Fabry disease who were treated in an open-label study with 1.0 mg/kg body weight of recombinant &agr;-Gal A (agalsidase &bgr;, Fabrazyme) were followed up for 12 months. Myocardial function was quantified by ultrasonic strain rate imaging to assess radial and longitudinal myocardial deformation. End-diastolic thickness of the left ventricular posterior wall and myocardial mass (assessed by magnetic resonance imaging, n=10) was measured at baseline and after 12 months of ERT. Data were compared with 16 age-matched healthy controls. At baseline, both peak systolic strain rate and systolic strain were significantly reduced in the radial and longitudinal direction in patients compared with controls. Peak systolic strain rate increased significantly in the posterior wall (radial function) after one year of treatment (baseline, 2.8±0.2 s−1; 12 months, 3.7±0.3 s−1; P <0.05). In addition, end-systolic strain of the posterior wall increased significantly (baseline, 34±3%; 12 months, 45±4%; P <0.05). This enhancement in radial function was accompanied by an improvement in longitudinal function. End-diastolic thickness of the posterior wall decreased significantly after 12 months of treatment (baseline, 13.8±0.6 mm; 12 months, 11.8±0.6 mm; P <0.05). In parallel, myocardial mass decreased significantly from 201±18 to 180±21 g (P <0.05). Conclusions—These results suggest that ERT can decrease left ventricular hypertrophy and improve regional myocardial function.

Endothelial Nitric Oxide Synthase Uncoupling Impairs Endothelial Progenitor Cell Mobilization and Function in Diabetes

Uncoupling of the endothelial nitric oxide synthase (eNOS) resulting in superoxide anion (O2−) formation instead of nitric oxide (NO) causes diabetic endothelial dysfunction. eNOS regulates mobilization and function of endothelial progenitor cells (EPCs), key regulators of vascular repair. We postulate a role of eNOS uncoupling for reduced number and function of EPC in diabetes. EPC levels in diabetic patients were significantly reduced compared with those of control subjects. EPCs from diabetic patients produced excessive O2− and showed impaired migratory capacity compared with nondiabetic control subjects. NOS inhibition with NG-nitro-l-arginine attenuated O2− production and normalized functional capacity of EPCs from diabetic patients. Glucose-mediated EPC dysfunction was protein kinase C dependent, associated with reduced intracellular BH4 (tetrahydrobiopterin) concentrations, and reversible after exogenous BH4 treatment. Activation of NADPH oxidases played an additional but minor role in glucose-mediated EPC dysfunction. In rats with streptozotocin-induced diabetes, circulating EPCs were reduced to 39 ± 5% of controls and associated with uncoupled eNOS in bone marrow. Our results identify uncoupling of eNOS in diabetic bone marrow, glucose-treated EPCs, and EPCs from diabetic patients resulting in eNOS-mediated O2− production. Subsequent reduction of EPC levels and impairment of EPC function likely contributes to the pathogenesis of vascular disease in diabetes.

Direct evidence for a β1-adrenergic receptor–directed autoimmune attack as a cause of idiopathic dilated cardiomyopathy

Today, dilated cardiomyopathy (DCM) represents the main cause of severe heart failure and disability in younger adults and thus is a challenge for public health. About 30% of DCM cases are genetic in origin; however, the large majority of cases are sporadic, and a viral or immune pathogenesis is suspected. Following the established postulates for pathogenesis of autoimmune diseases, here we provide direct evidence that an autoimmune attack directed against the cardiac β1-adrenergic receptor may play a causal role in DCM. First, we immunized inbred rats against the second extracellular β1-receptor loop (β1-ECII; 100% sequence identity between human and rat) every month. All these rats developed first, receptor-stimulating anti–β1-ECII Ab’s and then, after 9 months, progressive severe left ventricular dilatation and dysfunction. Second, we transferred sera from anti–β1-ECII–positive and Ab-negative animals every month to healthy rats of the same strain. Strikingly, all anti–β1-ECII–transferred rats also developed a similar cardiomyopathic phenotype within a similar time frame, underlining the pathogenic potential of these receptor Ab’s. As a consequence, β1-adrenergic receptor–targeted autoimmune DCM should now be categorized with other known receptor Ab-mediated autoimmune diseases, such as Graves disease or myasthenia gravis. Although carried out in an experimental animal model, our findings should further encourage the development of therapeutic strategies that combat harmful anti–β1-ECII in receptor Ab–positive DCM patients.

Beneficial effects of long-term intravenous iron therapy with ferric carboxymaltose in patients with symptomatic heart failure and iron deficiency

Aim The aim of this study was to evaluate the benefits and safety of long-term i.v. iron therapy in iron-deficient patients with heart failure (HF). Methods and results CONFIRM-HF was a multi-centre, double-blind, placebo-controlled trial that enrolled 304 ambulatory symptomatic HF patients with left ventricular ejection fraction ≤45%, elevated natriuretic peptides, and iron deficiency (ferritin <100 ng/mL or 100–300 ng/mL if transferrin saturation <20%). Patients were randomized 1 : 1 to treatment with i.v. iron, as ferric carboxymaltose (FCM, n = 152) or placebo (saline, n = 152) for 52 weeks. The primary end-point was the change in 6-min-walk-test (6MWT) distance from baseline to Week 24. Secondary end-points included changes in New York Heart Association (NYHA) class, Patient Global Assessment (PGA), 6MWT distance, health-related quality of life (QoL), Fatigue Score at Weeks 6, 12, 24, 36, and 52 and the effect of FCM on the rate of hospitalization for worsening HF. Treatment with FCM significantly prolonged 6MWT distance at Week 24 (difference FCM vs. placebo: 33 ± 11 m, P = 0.002). The treatment effect of FCM was consistent in all subgroups and was sustained to Week 52 (difference FCM vs. placebo: 36 ± 11 m, P < 0.001). Throughout the study, an improvement in NYHA class, PGA, QoL, and Fatigue Score in patients treated with FCM was detected with statistical significance observed from Week 24 onwards. Treatment with FCM was associated with a significant reduction in the risk of hospitalizations for worsening HF [hazard ratio (95% confidence interval): 0.39 (0.19–0.82), P = 0.009]. The number of deaths (FCM: 12, placebo: 14 deaths) and the incidence of adverse events were comparable between both groups. Conclusion Treatment of symptomatic, iron-deficient HF patients with FCM over a 1-year period resulted in sustainable improvement in functional capacity, symptoms, and QoL and may be associated with risk reduction of hospitalization for worsening HF (ClinicalTrials.gov number NCT01453608).

Long-Term Effects of Enzyme Replacement Therapy on Fabry Cardiomyopathy Evidence for a Better Outcome With Early Treatment

Background— Enzyme replacement therapy with recombinant α-galactosidase A reduces left ventricular hypertrophy and improves regional myocardial function in patients with Fabry disease during short-term treatment. Whether enzyme replacement therapy is effective in all stages of Fabry cardiomyopathy during long-term follow-up is unknown. Methods and Results— We studied 32 Fabry patients over a period of 3 years regarding disease progression and clinical outcome under enzyme replacement therapy. Regional myocardial fibrosis was assessed by magnetic resonance imaging late-enhancement technique. Echocardiographic myocardial mass was calculated with the Devereux formula, and myocardial function was quantified by ultrasonic strain-rate imaging. In addition, exercise capacity was measured by bicycle stress test. All measurements were repeated at yearly intervals. At baseline, 9 patients demonstrated at least 2 fibrotic left ventricular segments (severe myocardial fibrosis), 11 had 1 left ventricular segment affected (mild fibrosis), and 12 were without fibrosis. In patients without fibrosis, enzyme replacement therapy resulted in a significant reduction in left ventricular mass (238±42 g at baseline, 202±46 g at 3 years; P for trend <0.001), an improvement in myocardial function (systolic radial strain rate, 2.3±0.4 and 2.9±0.6 seconds−1, respectively; P for trend=0.045), and a higher exercise capacity obtained by bicycle stress exercise (106±14 and 122±26 W, respectively; P for trend=0.014). In contrast, patients with mild or severe fibrosis showed a minor reduction in left ventricular hypertrophy and no improvement in myocardial function or exercise capacity. Conclusions— These data suggest that treatment of Fabry cardiomyopathy with recombinant α-galactosidase A should best be started before myocardial fibrosis has developed to achieve long-term improvement in myocardial morphology and function and exercise capacity.

MicroRNA-24 Regulates Vascularity After Myocardial Infarction

Background— Myocardial infarction leads to cardiac remodeling and development of heart failure. Insufficient myocardial capillary density after myocardial infarction has been identified as a critical event in this process, although the underlying mechanisms of cardiac angiogenesis are mechanistically not well understood. Methods and Results— Here, we show that the small noncoding RNA microRNA-24 (miR-24) is enriched in cardiac endothelial cells and considerably upregulated after cardiac ischemia. MiR-24 induces endothelial cell apoptosis, abolishes endothelial capillary network formation on Matrigel, and inhibits cell sprouting from endothelial spheroids. These effects are mediated through targeting of the endothelium-enriched transcription factor GATA2 and the p21-activated kinase PAK4, which were identified by bioinformatic predictions and validated by luciferase gene reporter assays. Respective downstream signaling cascades involving phosphorylated BAD (Bcl-XL/Bcl-2–associated death promoter) and Sirtuin1 were identified by transcriptome, protein arrays, and chromatin immunoprecipitation analyses. Overexpression of miR-24 or silencing of its targets significantly impaired angiogenesis in zebrafish embryos. Blocking of endothelial miR-24 limited myocardial infarct size of mice via prevention of endothelial apoptosis and enhancement of vascularity, which led to preserved cardiac function and survival. Conclusions— Our findings indicate that miR-24 acts as a critical regulator of endothelial cell apoptosis and angiogenesis and is suitable for therapeutic intervention in the setting of ischemic heart disease.