Deficiency of MAPK-activated protein kinase 2 (MK2) prevents adverse remodelling and promotes endothelial healing after arterial injury.

Maladaptive remodelling of the arterial wall after mechanical injury (e. g. angioplasty) is characterised by inflammation, neointima formation and media hypertrophy, resulting in narrowing of the affected artery. Moreover, mechanical injury of the arterial wall causes loss of the vessel protecting endothelial cell monolayer. Mitogen-activated protein kinase (MAPK)-activated protein kinase 2 (MK2), a major downstream target of p38 MAPK, regulates inflammation, cell migration and proliferation, essential processes for vascular remodelling and re-endothelialisation. Therefore, we investigated the role of MK2 in remodelling and re-endothelialisation after arterial injury in genetically modified mice in vivo. Hypercholesterolaemic low-density-lipoprotein-receptor-deficient mice (ldlr-/-) were subjected to wire injury of the common carotid artery. MK2-deficiency (ldlr-/-/mk2-/-) nearly completely prevented neointima formation, media hypertrophy, and lumen loss after injury. This was accompanied by reduced proliferation and migration of MK2-deficient smooth muscle cells. In addition, MK2-deficiency severely reduced monocyte adhesion to the arterial wall (day 3 after injury, intravital microscopy), which may be attributed to reduced expression of the chemokine ligands CCL2 and CCL5. In line, MK2-deficiency significantly reduced the content of monocytes, neutrophiles and lymphocytes of the arterial wall (day 7 after injury, flow cytometry). In conclusion, in a model of endothelial injury (electric injury), MK2-deficiency strongly increased proliferation of endothelial cells and improved re-endothelialisation of the arterial wall after injury. Deficiency of MK2 prevents adverse remodelling and promotes endothelial healing of the arterial wall after injury, suggesting that MK2-inhibition is a very attractive intervention to prevent restenosis after percutaneous therapeutic angioplasty.

Diagnostic and prognostic performance of a novel high-sensitivity cardiac troponin T assay compared to a contemporary sensitive cardiac troponin I assay in patients with acute coronary syndrome.

OBJECTIVE: The study sought to compare the clinical performance of two more sensitive cardiac troponin (cTn) assays, a novel high-sensitivity (hs) troponin T assay and a contemporary cTnI assay. METHODS: We measured hs-cTnT (Roche TnThs) and cTnI (Siemens Centaur Ultra) on presentation in 1,384 patients with suspected acute coronary syndrome (ACS) who underwent early invasive strategy within 24 h after presentation. Kaplan-Meier, Cox proportional hazards, and receiver-operating characteristic (ROC) analysis was used to compare their prognostic performance for the prediction of all-cause death and death/MI (myocardial infarction) after a median of 271 days. We also compared the diagnostic performance of these assays on presentation for early diagnosis of non-STEMI. RESULTS: Both hs-cTnT and cTnI were independently predictive of long-term death (OR 3.51 vs. 2.19) and the composite of death/MI (OR 9.24 vs. 3.61), across the spectrum of ACS and in patients without ACS. When used as a continuous variable, ROC analysis demonstrated significantly higher areas under the curve (AUC) for hs-cTnT as compared to cTnI for the prediction of death/MI (0.721 vs. 0.672, P = 0.024), a trend to better prediction of all-cause death (0.721 vs. 0.672, P = 0.093) and significantly higher AUC for early diagnosis of non-STEMI (0.965 vs. 0.901, P < 0.001). CONCLUSION: Using the 99th percentile cutoff for hs-cTnT and cTnI, both assays enable prediction of adverse long-term outcomes and earlier diagnosis of non-STEMI. Used as a continuous variable, the hs-cTnT assay showed superior performance compared to the cTnI assay, especially in regard to prognosis.

Cardiac troponin T concentrations above the 99th percentile value as measured by a new high-sensitivity assay predict long-term prognosis in patients with acute coronary syndromes undergoing routine early invasive strategy.

OBJECTIVE: A recently developed immunoassay for high-sensitivity measurement of cardiac troponin T (hsTnT) allows measurement at the 99th percentile for a normal population with an assay imprecision <10%. It is unclear whether such a low cutpoint (14 ng/L) is helpful for long-term risk stratification of patients with an acute coronary syndrome (ACS) undergoing routine early invasive strategy. PATIENTS AND MAIN OUTCOME MEASURES: Consecutive patients with ACS admitted to a chest pain unit were studied. The usefulness of hsTnT for early diagnosis of myocardial infarction (MI) and prediction of all-cause death or death/MI over a median of 271 days following presentation was compared against the fourth generation cTnT at the 99th percentile cutpoint. RESULTS: Of 1,384 patients with ACS enrolled, 47.8% had non-ST-segment elevation MI (NSTEMI), 26.4% unstable angina, 21.8% STEMI and 4% had non-ACS. Adjusted risk for all-cause death [adjusted HR 8.26 (95%CI: 1.13-66.33), p = 0.038] and death/MI [adjusted HR 2.71 (95% CI: 1.15-6.38), p = 0.023] were significantly higher with hsTnT above the 99th percentile. In particular, among patients with a standard fourth generation cTnT result below the 99th percentile cutoff (0.01 ng/mL), hsTnT improved risk assessment. Mortality risk associated with an elevated hsTnT was present across the spectrum of ACS, as well as in conditions with hsTnT elevations not related to ACS. CONCLUSION: hsTnT at the 99th percentile cutoff is useful for the diagnostic evaluation of patients with ACS, and provides strong and independent predictive power for adverse long-term outcomes even after early invasive strategy.

A simple non-invasive diagnostic algorithm for ruling out chronic thromboembolic pulmonary hypertension in patients after acute pulmonary embolism.

BACKGROUND: Our aim was to construct a diagnostic model for ruling out chronic thromboembolic pulmonary hypertension (CTEPH) in symptomatic patients after acute pulmonary embolism (PE) that is based on simple, non-invasive tests. METHODS: Plasma levels of various CTEPH associated biomarkers and conventional ECG criteria for right ventricular pressure overload were assessed in 82 consecutive patients with confirmed CTEPH and 160 consecutive patients with a history of PE who were suspected to have CTEPH, but in whom this disease was ruled out. RESULTS: ECG criteria of right ventricular hypertrophy were detected more frequently in patients with CTEPH (77%) than in the patients without CTEPH (11%, Odds ratio 26, 95% confidence interval [CI] 13-53). Also, clotting factor FVIII activity and the levels of N-terminal-pro-brain-type natriuretic peptide (NT-pro-BNP), Growth Differentiation Factor-15, C-reactive protein and urate, but not D-dimer level, were higher in patients with CTEPH. A diagnostic model including ECG criteria and NT-pro-BNP levels had a sensitivity of 94% (95% CI 86-98%) and a specificity of 65% (95% CI 56-72%). The area under the receiver-operator-characteristic curve was 0.80 (95% CI 0.74-0.85) for the diagnosis of CTEPH. Even with high disease prevalences of up to 10%, the negative predictive value of this model proved to be very high (99%, 95% CI 97-100%). CONCLUSIONS: Ruling out CTEPH in patients after acute PE seems to be safe without additional diagnostic testing in absence of ECG criteria indicative of right ventricular hypertrophy and a normal NT-pro-BNP level.

[Pathophysiology of heart failure]. / Pathophysiologie der Herzinsuffizienz.

Chronic heart failure is a clinical syndrome and the final common pathway of different cardiac diseases. Heart failure is accompanied by activation of the renin-angiotensin-aldosterone-system and the adrenergic nervous system. In addition, recent data emphasize important roles of maladaptive intracellular signaling pathways, decreased capillary density, altered calcium handling, metabolic changes, genetic polymorphisms, and programmed cell death in the failing heart. In this context, traditional pathophysiological concepts, e. g. concerning the role of cardiac hypertrophy, had to be given up. Thus, an increasingly complex scenario emerges with interdependent changes on the biochemical, molecular, metabolic, and cellular level. Novel therapeutic strategies may soon be based on these new pathophysiological concepts.

Good manufacturing practice-compliant validation and preparation of BM cells for the therapy of acute myocardial infarction.

BACKGROUND: Intracoronary application of BM-derived cells for the treatment of acute myocardial infarction (AMI) is currently being studied intensively. Simultaneously, strict legal requirements surround the production of cells for clinical studies. Thus good manufacturing practice (GMP)-compliant collection and preparation of BM for patients with AMI was established by the Cytonet group. METHODS: As well as fulfillment of standard GMP requirements, including a manufacturing license, validation of the preparation process and the final product was performed. Whole blood (n=6) and BM (n=3) validation samples were processed under GMP conditions by gelafundin or hydroxyethylstarch sedimentation in order to reduce erythrocytes/platelets and volume and to achieve specifications defined in advance. Special attention was paid to the free potassium (<6 mmol/L), some rheologically relevant cellular characteristics (hematocrit <0.45, platelets <450 x 10(6)/mL) and the sterility of the final product. RESULTS: The data were reviewed and GMP compliance was confirmed by the German authorities (Paul-Ehrlich Institute). Forty-five BM cell preparations for clinical use were carried out following the validated methodology and standards. Additionally three selections of CD34+ BM cells for infusion were performed. All specification limits were met. Discussion In conclusion, preparation of BM cells for intracoronary application is feasible under GMP conditions. As the results of sterility testing may not be available at the time of intracoronary application, the highest possible standards to avoid bacterial and other contaminations have to be applied. The increased expense of the GMP-compliant process can be justified by higher safety for patients and better control of the final product.

Stem cell therapy: a new perspective in the treatment of patients with acute myocardial infarction.

Experimental studies suggest that cardiac transfer of stem and progenitor cells can have a favorable impact on tissue perfusion and contractile performance after acute myocardial infarction (AMI). While the mechanistic background of stem cell therapy is still intensely debated, the concept of cell therapy has already been introduced into the clinical setting, where small, mostly uncontrolled trials indicate that stem cell therapy may be feasible in patients. The overall clinical experience also suggests that stem cell therapy can be safely performed, if the right cell type is used in the right clinical setting. Preliminary efficacy data indicate that stem cells have the potential to enhance myocardial perfusion and/or contractile performance in patients with AMI. The field now is rapidly moving towards intermediate-size, double-blinded trials to gather more safety and efficacy data. Ultimately, large outcome trials will have to be conducted. At the same time, continued basic research to elucidate the underlying mechanism of stem cell therapy is needed.

Euro heart failure survey. Medical treatment not in line with current guidelines.

UNLABELLED: It was the aim of the Euro Heart Survey on Heart Failure to assess whether patients are being treated according to current guidelines. METHODS: In Germany, patients were screened in 7 medical centers if their discharge diagnoses were myocardial infarction, a new episode of atrial fibrillation, or diabetes mellitus. Patients were enrolled if at least one additional criterion was fulfilled: (1) clinical diagnosis of heart failure, (2) hospital admission due to heart failure within the last 3 years, (3) therapy with loop diuretic, (4) medication for heart failure or ventricular dysfunction documented by echocardiography within the past 24 hours prior to death. RESULTS: 2166 patients were screened of whom 747 were included in the study (478 men, 269 women). 93% of the patients suffered from heart failure. Despite the high number of patients with known heart failure (ischemic heart failure in 71%), only 72% received ACE inhibitors and 62% beta-blockers. Average daily dose met recommendations in only 63% of patients on ACE inhibitors and 54% on beta-blockers. 74% of the patients received diuretics (furosemide 36%, thiazide 34%, spironolactone 17%). CONCLUSION: An inadequately low number of patients with heart failure receives medical therapy according to guidelines, despite all the overwhelming evidence for improved morbidity and mortality. Awareness of physicians needs to be improved.

The role of interleukin-6 in the failing heart.

Clinical studies have shown that circulating levels of interleukin (IL)-6 and other IL-6 related cytokines are increased in patients with congestive heart failure (CHF). Plasma IL-6 concentrations are related to decreasing functional status of the patient and provide important prognostic information. Experimental studies have produced compelling evidence that IL-6 and IL-6 related cytokines play a pivotal role in the regulation of cardiac myocyte hypertrophy and apoptosis. This review summarizes clinical and experimental data from this rapidly evolving field, which, taken together, strongly suggest that IL-6 and IL-6 related cytokines are intricately involved in the pathophysiology of the failing heart.

A novel genetic pathway for sudden cardiac death via defects in the transition between ventricular and conduction system cell lineages.

HF-1 b, an SP1 -related transcription factor, is preferentially expressed in the cardiac conduction system and ventricular myocytes in the heart. Mice deficient for HF-1 b survive to term and exhibit normal cardiac structure and function but display sudden cardiac death and a complete penetrance of conduction system defects, including spontaneous ventricular tachycardia and a high incidence of AV block. Continuous electrocardiographic recordings clearly documented cardiac arrhythmogenesis as the cause of death. Single-cell analysis revealed an anatomic substrate for arrhythmogenesis, including a decrease and mislocalization of connexins and a marked increase in action potential heterogeneity. Two independent markers reveal defects in the formation of ventricular Purkinje fibers. These studies identify a novel genetic pathway for sudden cardiac death via defects in the transition between ventricular and conduction system cell lineages.

The cardiac Fas (APO-1/CD95) Receptor/Fas ligand system : relation to diastolic wall stress in volume-overload hypertrophy in vivo and activation of the transcription factor AP-1 in cardiac myocytes.

BACKGROUND: Fas (APO-1/CD95) is a transmembrane receptor belonging to the tumor necrosis factor receptor superfamily. Cross-linking of Fas by Fas ligand (FasL), a tumor necrosis factor-alpha-related cytokine, promotes apoptosis and/or transcription factor activation in a highly cell-type-specific manner. The biological consequences of Fas activation in cardiomyocytes and the regulation of Fas and FasL abundance in the myocardium in vivo remain largely unknown. METHODS AND RESULTS: As shown by immunohistochemistry, Fas was expressed on the sarcolemma of cardiomyocytes in left ventricular tissue sections. Moreover, FasL was constitutively expressed in the myocardium and in isolated cardiomyocytes, as revealed by reverse transcription polymerase chain reaction and Western blotting. Left ventricular abundance of Fas but not FasL was upregulated in a rat model of compensated volume-overload hypertrophy and was closely related to diastolic but not systolic wall stress as determined by MRI. Cardiomyocyte apoptosis was not enhanced in volume-overload hypertrophy despite the increased expression of Fas and the presence of FasL in the myocardium. Moreover, injection of mice with an agonistic anti-Fas antibody promoted hepatocyte but not cardiomyocyte apoptosis in vivo. Stimulation of isolated cardiomyocytes with recombinant FasL promoted an activation of the transcription factor AP-1 as shown by electrophoretic mobility shift assays but did not induce cell death. CONCLUSIONS: Fas and FasL are constitutively expressed in the myocardium and in cardiomyocytes. Myocardial expression of Fas is closely related to diastolic loading conditions in vivo. Signaling pathways emanating from Fas are coupled to an activation of the transcription factor AP-1 in cardiomyocytes.

The renin-angiotensin system and experimental heart failure.

Experimental studies suggest that the renin-angiotensin system (RAS) and its primary effector peptide, angiotensin II (Ang II), are involved in the pathophysiology of cardiac hypertrophy and failure. All the components required for Ang II production are present in the heart, and cardiac Ang II formation appears to be regulated independent from the circulating RAS. In animal models and in patients with heart failure, the cardiac RAS is activated and, presumably, local Ang II formation is enhanced. Several cardiac cell types express Ang II type 1 (AT1) and/or type 2 (AT2)-receptors and represent potential targets for Ang II-mediated effects. In neonatal cardiac myocytes, Ang II induces a hypertrophic response via the AT1-receptor. Likewise, activation of the AT1-receptor triggers hypertrophy in terminally differentiated cardiac myocytes and in perfused heart preparations. In the neonatal system, Ang II appears to be a major autocrine/paracrine mediator of cardiac myocyte hypertrophy in response to passive mechanical stretch. By contrast, AT1-receptor activation apparently is not required to trigger load-induced hypertrophy in the adult cardiomyocyte. Recent studies suggest that the AT2-receptor opposes AT1-receptor-mediated growth signals in neonatal and in adult cardiac myocytes. Pharmacological studies have established that a blockade of the RAS at the level of the angiotensin-converting enzyme (ACE) or the AT1-receptor ameliorates the remodeling process of the heart and prolongs long-term survival in animal models of cardiac hypertrophy and failure. The therapeutic effects of ACE inhibitors and AT1-receptor antagonists clearly suggest an important role for the ACE-Ang II-AT1-receptor axis in the development of cardiac hypertrophy and failure. It must be kept in mind, however, that these drugs enhance AT2-receptor and B2-kinin receptor-dependent signaling pathways which may contribute significantly to the beneficial effects observed in vivo. Molecular and physiological analyses of transgenic mice with a cardiac-specific overexpression of the AT1 or AT2-receptor confirm that AT1 and AT2-receptor-dependent signaling cascades potently modulate cardiac myocyte function and growth. However, studies in AT1-receptor knockout mice demonstrate that cardiac hypertrophy in response to hemodynamic overload can occur independent from the AT1-receptor. In this paper, we review recent experimental evidence suggesting a critical role for the RAS in cardiac hypertrophy and failure with special emphasis on the putative role of Ang II and Ang II-receptor signaling in cardiac myocytes.

The kallikrein-kinin system in post-myocardial infarction cardiac remodeling.

Angiotensin converting-enzyme (ACE) inhibitors attenuate cardiac hypertrophy and prolong survival in animal models and patients after myocardial infarction (MI). Considering the dual function of the ACE, the therapeutic efficacy of ACE inhibitors after MI implicates the renin-angiotensin system and/or the kallikrein-kinin system in the pathophysiology of postinfarction cardiac remodeling. We evaluated the role of kinins, and their potential contribution to the antiremodeling effects of ACE inhibition in this setting. Rats underwent coronary artery ligation followed by chronic B2 kinin receptor blockade with icatibant (HOE 140). Additional groups of MI rats were treated with the ACE inhibitor lisinopril, alone or in combination with icatibant. B2 kinin receptor blockade enhanced the deposition of collagen (morphometric analysis) in the left ventricular interstitial space after MI, whereas markers of cardiomyocyte hypertrophy (left ventricular weights and prepro-atrial natriuretic factor [ANF] expression) were not affected. Chronic ACE inhibition reduced collagen deposition and cardiomyocyte hypertrophy after MI. The inhibitory action of ACE inhibition on interstitial collagen was partially reversed by B2 kinin receptor blockade. However, B2 kinin receptor blockade did not attenuate the effects of ACE inhibition on cardiomyocyte hypertrophy. In conclusion, kinins inhibit the interstitial accumulation of collagen, but do not modulate cardiomyocyte hypertrophy after MI. Kinins contribute to the reduction of myocardial collagen accumulation by ACE inhibition; however, the effects of ACE inhibition on cardiomyocyte hypertrophy are related to reduced generation of angiotensin II.

Cardiotrophin-1 and the role of gp130-dependent signaling pathways in cardiac growth and development.

The reactivation of an embryonic pattern of gene expression is a central feature common to virtually all forms of cardiac hypertrophy. Unraveling the regulatory mechanisms, growth factors and cytokines controlling gene expression and cell fate during cardiac development may therefore have implications for our understanding of cardiac hypertrophy in the adult. Along this line, a cDNA expression library was established from an embryonic stem cell-based in vitro model of cardiogenesis, and screened for clones that would induce an increase in cell size in cultured cardiomyocytes. This experimental strategy resulted in the isolation of a novel cytokine, cardiotrophin-1 (CT-1), that activates several features of cardiomyocyte hypertrophy in vitro, including sarcomeric organization and embryonic gene expression. CT-1 displays structural similarities to the interleukin (IL)-6 related cytokines. Furthermore, receptor binding studies and functional studies reveal that CT-1 shares the signal transducing receptor components gp130 and LIFR with the previously identified members of the IL-6 cytokine family. CT-1 rapidly activates gp130 and LIFR tyrosine phosphorylation in cultured cardiac myocytes. The growth promoting effects of CT-1 therefore indicate that signaling pathways emanating from gp130 and LIFR are coupled to cardiomyocyte hypertrophy. In support of this notion, the simultaneous overexpression of IL-6 and the IL-6 receptor in transgenic mice has been shown to result in a constitutive tyrosine phosphorylation of gp130 in the myocardium and cardiac hypertrophy. The striking phenotype of gp130 null-mutant mice, generated by homologous recombination, implies gp130 in cardiac development as well: mutant mice exhibit severe ventricular hypoplasia, suggesting a role for gp130-dependent signaling pathways in the expansion of the compact layer of the ventricular myocardium. CT-1 is expressed at high levels in the myocardium during the course of cardiogenesis, and promotes the proliferation and survival of embryonic cardiomyocytes. CT-1 may therefore represent a candidate cytokine to activate gp130 during cardiac development. In summary, cytokines signaling through gp130 are emerging as potent regulators of embryonic heart development and adult cardiac hypertrophy.

Differential effects of kinins on cardiomyocyte hypertrophy and interstitial collagen matrix in the surviving myocardium after myocardial infarction in the rat.

BACKGROUND: Left ventricular remodeling after myocardial infarction (MI) involves the hypertrophic growth of cardiomyocytes and the accumulation of fibrillar collagen in the interstitial space. We evaluated the role of kinins in postinfarction ventricular remodeling and their potential contribution to the antiremodeling effects of ACE inhibition and angiotensin II type 1 (AT1) receptor blockade. METHODS AND RESULTS: Rats underwent coronary artery ligation followed by chronic B2 kinin receptor blockade with icatibant. Additional groups of infarcted rats were treated with the ACE inhibitor lisinopril or the AT1 receptor antagonist ZD7155, each separately and in combination with icatibant. B2 kinin receptor blockade enhanced the interstitial deposition of collagen after MI, whereas morphological and molecular markers of cardiomyocyte hypertrophy (cardiac weight, myocyte cross-sectional area, prepro-atrial natriuretic factor mRNA expression) were not affected. Chronic ACE inhibition and AT1 receptor blockade reduced collagen deposition and cardiomyocyte hypertrophy after MI. The inhibitory action of ACE inhibition and AT1 receptor blockade on interstitial collagen was partially reversed by B2 kinin receptor blockade. However, B2 kinin receptor blockade did not attenuate the effects of ACE inhibition and AT1 receptor blockade on cardiomyocyte hypertrophy. CONCLUSIONS: (1) Kinins inhibit the interstitial accumulation of collagen but do not modulate cardiomyocyte hypertrophy after MI. (2) Kinins contribute to the reduction of myocardial collagen accumulation by ACE inhibition and AT1 receptor blockade. (3) The effects of ACE inhibition and AT1 receptor blockade on cardiomyocyte hypertrophy are related to a reduced generation/receptor blockade of angiotensin II.

Cardiotrophin-1 activates a distinct form of cardiac muscle cell hypertrophy. Assembly of sarcomeric units in series VIA gp130/leukemia inhibitory factor receptor-dependent pathways.

Cardiotrophin-1 (CT-1) was recently isolated by expression cloning based on its ability to induce an increase in cell size in neonatal rat ventricular cardiomyocytes. Sequence similarity data suggested that CT-1 is a novel member of a family of structurally related cytokines sharing the receptor component gp130. The present study documents that gp130 is required for CT-1 signaling in cardiomyocytes, by demonstrating that a monoclonal anti-gp130 antibody completely inhibits c-fos induction by CT-1. Similarly, a leukemia inhibitory factor receptor subunit beta (LIFRbeta) antagonist effectively blocks the CT-1 induction of c-fos, indicating a requirement for LIFRbeta in the hypertrophic response, as well. Upon stimulation with CT-1, both gpl30 and the LIFRbeta are tyrosine-phosphorylated, providing further evidence that CT-1 signals through the gp130/LIFRbeta heterodimer in cardiomyocytes. CT-1 induces a hypertrophic response in cardiomyocytes that is distinct from the phenotype seen after alpha-adrenergic stimulation, both with regard to cell morphology and gene expression pattern. Stimulation with CT-1 results in an increase in cardiac cell size that is characterized by an increase in cell length but no significant change in cell width. Confocal laser microscopy of CT-1 stimulated cells reveals the assembly of sarcomeric units in series rather than in parallel, as seen after alpha-adrenergic stimulation. CT-1 induces a distinct pattern of immediate early genes, and up-regulates the atrial natriuretic factor (ANF) gene, but does not affect skeletal alpha-actin or myosin light chain-2v expression. As evidenced by nuclear run-on transcription assays, both CT-1 and alpha-adrenergic stimulation lead to an increase in ANF gene transcription. Transient transfection analyses document that, in contrast to alpha-adrenergic stimulation, the CT-1 responsive cis-regulatory elements are located outside of the proximal 3 kilobase pairs of the ANF 5'-flanking region. These studies indicate that CT-1 can activate a distinct form of myocardial cell hypertrophy, characterized by the promotion of sarcomere assembly in series, via gpl30/LIFRbeta-dependent signaling pathways.