Correlation of gene expression and clinical parameters identifies a set of genes reflecting LV systolic dysfunction and morphological alterations.

To gain new insights into the complex pathophysiology of dilated cardiomyopathy (DCM) we performed a quantitative approach to identify genes with expression patterns that linearly correlate with parameters of cardiac morphology (left ventricular end-diastolic diameter indexed by body surface are (LVEDDI), systolic function [LV ejection fraction (LVEF)], and serum levels of cardiac peptide hormone NH2-terminal probrain natriuretic peptide (NT-proBNP) in human endomyocardial biopsies of 47 DCM patients and eight individuals with normal LVEF. A set of genes was identified as common heart failure markers characterized by correlation of their expression with cardiac morphology, systolic function, and NT-proBNP. Among them are already known genes encoding e.g., the natriuretic peptide hormones NPPA and NPPB and its converting enzyme corin, but also potential new heart failure markers like EP300 antisense RNA1 and dimethylarginine dimethylaminohydrolase 1 (DDAH1) along with other genes with so far unknown relation to heart function. In contrast, the expression of other genes including the Ca2+ flux regulating genes phospholamban (PLN), sarcoplasmic/endoplasmic reticulum calcium ATPase 2 (SERCA), and extracellular matrix proteins showed significant correlation with LVEF and LVEDDI only. Those genes seem to reflect more specifically pathological alterations of systolic function and morphology in DCM hearts.

Changes of myocardial gene expression and protein composition in patients with dilated cardiomyopathy after immunoadsorption with subsequent immunoglobulin substitution.

Immunoadsorption with subsequent immunoglobulin substitution (IA/IgG) represents a therapeutic approach for patients with dilated cardiomyopathy (DCM). Here, we studied which molecular cardiac alterations are initiated after this treatment. Transcription profiling of endomyocardial biopsies with Affymetrix whole genome arrays was performed on 33 paired samples of DCM patients collected before and 6 months after IA/IgG. Therapy-related effects on myocardial protein levels were analysed by label-free proteome profiling for a subset of 23 DCM patients. Data were analysed regarding therapy-associated differences in gene expression and protein levels by comparing responders (defined by improvement of left ventricular ejection fraction ≥20 % relative and ≥5 % absolute) and non-responders. Responders to IA/IgG showed a decrease in serum N-terminal proBNP levels in comparison with baseline which was accompanied by a decreased expression of heart failure markers, such as angiotensin converting enzyme 2 or periostin. However, despite clinical improvement even in responders, IA/IgG did not trigger general inversion of DCM-associated molecular alterations in myocardial tissue. Transcriptome profiling revealed reduced gene expression for connective tissue growth factor, fibronectin, and collagen type I in responders. In contrast, in non-responders after IA/IgG, fibrosis-associated genes and proteins showed elevated levels, whereas values were reduced or maintained in responders. Thus, improvement of LV function after IA/IgG seems to be related to a reduced gene expression of heart failure markers and pro-fibrotic molecules as well as reduced fibrosis progression.

Analysis of DCM associated protein alterations of human right and left ventricles.

Dilated cardiomyopathy (DCM) is characterized by ventricular chamber enlargement and impaired myocardial function. Endomyocardial biopsies (EMB) enable immunohistochemical and molecular characterization of this disease. However, knowledge about specific molecular patterns and their relation to cardiac function in both ventricles is rare. Therefore, we performed a mass spectrometric analysis of 28 paired EMBs of left (LV) and right ventricles (RV) of patients with DCM or suspected myocarditis allowing quantitative profiling of 743 proteins. We analysed associations between protein abundance of LV and RV as well as the echocardiographic parameters LVEF, TAPSE, LVEDDI, and RVEDDI by linear regression models. Overall, more LV than RV proteins were associated with LV parameters or with RVEDDI. Most LV and RV proteins increasing in level with impairing of LVEF were annotated to structural components of cardiac tissue. Additionally, a high proportion of LV proteins with metabolic functions decreased in level with decreasing LVEF. Results were validated with LV heart sections of a genetic murine heart failure model. The study shows, that remodelling and systolic dysfunction in DCM is mirrored by distinct alterations in protein composition of both ventricles. Loss of LV systolic function is reflected predominantly by alterations in proteins assigned to metabolic functions in the LV whereas structural remodelling was more obvious in the RV. Alterations related to intermediate filaments were seen in both ventricles and highlight such proteins as early indicators of LV loss of function. SIGNIFICANCE: The present study report protein sets in the RV and the LV being associated with ventricular function and remodelling in DCM. Protein abundances in the LV and the RV emphasize and expand current knowledge on pathophysiological changes in heart failure and DCM. While RV and LV EMBs do not differ concerning diagnostic assessment of inflammatory status and virus persistence, additional information reflecting disease severity associated protein alterations can be gained by EMB protein profiling. RV and LV protein data provided complementary information. The protein pattern of the LV reflects metabolic changes and an impaired energy production, which is associated with the degree of LV systolic dysfunction and remodelling and may yield important information about the disease status in DCM. On the other hand, at this disease stage of DCM with still preserved RV function, RV alterations in structural proteins may reflect myocardial compensatory protective mechanisms for maintenance of structure and cellular function. The study highlight particular proteins being of interest as heart failure biomarkers in both ventricles which seem to reflect the severity of the disease. Further comparative studies between different HF aetiologies have to evaluate those proteins as markers specific for DCM.

Effects of immunoadsorption and subsequent immunoglobulin G substitution on cardiopulmonary exercise capacity in patients with dilated cardiomyopathy.

BACKGROUND: Recent data indicate that cardiac antibodies play an active role in the pathogenesis of dilated cardiomyopathy (DCM) and may contribute to cardiac dysfunction in patients with DCM. The present study investigated the influence of immunoadsorption with subsequent immunoglobulin G substitution (IA/IgG) on cardiopulmonary exercise capacity in patients with DCM. METHODS: Sixty patients with DCM (New York Heart Association II-IV, left ventricular ejection fraction < or =45%) were included in this single-center university hospital-based case-control study. Patients either were treated with IA/IgG (n = 30) or were followed without IA/IgG (n = 30). At baseline and after 3 months, we compared echocardiographic assessment of left ventricular function and spiroergometric exercise parameters. RESULTS: In contrast to controls, left ventricular ejection fraction improved significantly in the IA/IgG group from 33.0% +/- 1.2% to 40.1% +/- 1.5% (P < .001). In the control group, spiroergometric exercise parameters did not change during follow-up. After 3 months, maximum achieved power increased in the treatment group from 114.2 +/- 7.4 to 141.9 +/- 7.9 W (P = .02). Total exercise time increased in the treatment group from 812 +/- 29 to 919 +/- 30 seconds (P < .05). Peak oxygen uptake (Vo(2)) increased from 17.3 +/- 0.9 to 21.8 +/- 1.0 mL min(-1) kg(-1) after IA/IgG (P < .01). Oxygen pulse (peak Vo(2)/maximum heart rate) increased in the treatment group (10.7 +/- 0.7 vs 13.6 +/- 0.7 mL beat(-1) min(-1), P < .01). The Vo(2) at the gas exchange anaerobic threshold increased after 3 months in the treatment group from 10.3 +/- 0.5 to 13.2 +/- 0.5 mL min(-1) kg(-1) (P < .001). The ventilatory response to exercise (V(E)/Vco(2) slope) decreased after IA/IgG therapy from 32.3 +/- 1.5 to 28.7 +/- 0.9 (P = .02). CONCLUSIONS: In patients with DCM, IA/IgG therapy may induce improvement in echocardiographic and cardiopulmonary exercise parameters.

[Update Myocarditis]. / Update Myokarditis.

Myocarditis is a heterogenous disease regarding aetiology, clinical presentation and course. A defined diagnostic procedure is needed to reliably detect myocarditis. While findings from medical laboratory parameters, electrocardiography and echocardiography are rather unspecific, endomyocardial biopsies supply dependable data regarding inflammatory and viral status. Analysis of cardiac MR is constantly being improved in order to increase sensitivity regarding myocarditis detection. Incidence, clinical presentation as well as disease progression and prognosis show considerable gender differences. Adamant implementation of defined diagnostic procedures is needed in order not to overlook myocarditis in women while the understanding of the pathomechanisms behind the gender differences might lead the way to new therapeutic options. Currently, treatment of myocarditis symptoms and heart failure is in the focus of clinical care. In addition, cardiac involvement in systemic inflammatory diseases should be stringently treated via immunosuppression. Avoidance of physical exercise has to be observed in order to reduce cardiac strain and consequently the number of adverse events.

Global plasma protein profiling reveals DCM characteristic protein signatures.

To identify potential biomarkers supporting better phenotyping and to improve understanding of the pathophysiology of dilated cardiomyopathy (DCM), this study comparatively analyzed plasma protein profiles of DCM patients and individuals with low normal and normal left ventricular ejection fraction (LVEF) by mass spectrometry. After plasma depletion using a MARS Hu-6 column, global proteome profiling was performed using a LTQ-Orbitrap Velos mass spectrometer. To compare and confirm results, two different discovery sets of samples were investigated. Differentially abundant proteins are involved in lipid metabolism, coagulation, and acute phase response. Serum paraoxonase 1 (PON1), cystatin C, lysozyme C, apolipoprotein A-II, and apolipoprotein M were validated by targeted protein analysis in a third independent patient cohort. Additionally, PON1 levels were also determined by an ELISA. These data highlight PON1 as a potential marker for differentiating DCM patients not only from patients with normal LVEF, but also from heart failure patients with preserved ejection fraction. The results highlight lipid metabolism and inflammation as the major pathways being altered in DCM patients in comparison to patients presenting with suspicious myocarditis to the hospital. SIGNIFICANCE: Several studies focused on the identification of heart failure (HF) associated protein signatures in blood plasma, but only few that are largely based on only small sample series considered specific HF pathologies. Therefore, we performed a comparative global blood plasma protein profiling of a larger sample of individuals with reduced left ventricular ejection fraction (LVEF) classified as dilated cardiomyopathy patients and individuals with normal LVEF but presenting with suspicious myocarditis. DCM patients displayed altered levels of proteins involved in lipid metabolism, coagulation, and acute phase response. The most reliable candidates, such as serum paraoxonase 1 (PON1), cystatin C, lysozyme C, apolipoprotein A-II, and apolipoprotein M were validated by targeted protein analysis in an independent patient cohort. PON1 levels were also determined by an ELISA. These data highlight PON1 as a potential marker for differentiating DCM patients not only from patients with normal LVEF, but also from heart failure patients with preserved ejection fraction.

Clinical Predictors and Prognostic Impact of Recovery of Wall Motion Abnormalities in Takotsubo Syndrome: Results From the International Takotsubo Registry.

Background Left ventricular (LV) recovery in takotsubo syndrome (TTS) occurs over a wide-ranging interval, varying from hours to weeks. We sought to investigate the clinical predictors and prognostic impact of recovery time for TTS patients. Methods and Results TTS patients from the International Takotsubo Registry were included in this study. Cut-off for early LV recovery was determined to be 10 days after the acute event. Multivariable logistic regression was used to assess factors associated with the absence of early recovery. In-hospital outcomes and 1-year mortality were compared for patients with versus without early recovery. We analyzed 406 patients with comprehensive and serial imaging data regarding time to recovery. Of these, 191 (47.0%) had early LV recovery and 215 (53.0%) demonstrated late LV improvement. Patients without early recovery were more often male (12.6% versus 5.2%; P=0.011) and presented more frequently with typical TTS (76.3% versus 67.0%, P=0.040). Cardiac and inflammatory markers were higher in patients without early recovery than in those with early recovery. Patients without early recovery showed unfavorable 1-year outcome compared with patients with early recovery (P=0.003). On multiple logistic regression, male sex, LV ejection fraction <45%, and acute neurologic disorders were associated with the absence of early recovery. Conclusions TTS patients without early LV recovery have different clinical characteristics and less favorable 1-year outcome compared with patients with early recovery. The factors associated with the absence of early recovery included male sex, reduced LV ejection fraction, and acute neurologic events. Clinical Trial Registration URL: http://www.clinicaltrials.gov. Unique identifier: NCT01947621.

MD-2 is a new predictive biomarker in dilated cardiomyopathy and exerts direct effects in isolated cardiomyocytes.

BACKGROUND: Myeloid differentiation factor-2 (MD-2) has been shown to be an important modulator of the innate immune system, but its role in cardiac diseases is unknown. We investigated whether MD-2 plays a role as risk predictor and contributor in dilated cardiomyopathy (DCM). METHODS AND RESULTS: We included 174 patients with reduced left ventricular (LV) ejection fraction (LVEF <45%) due to DCM. Coronary artery disease and severe valvular diseases were excluded in all patients by angiography or echocardiography. Cardiac inflammation, viral infection and MD-2 expression were analyzed from right ventricular endomyocardial biopsies. MD-2 was quantified by ELISA in serum upon first hospital admission. Myocyte contractility and inflammatory response after stimulation with recombinant MD-2 protein were analyzed in isolated rat cardiomyocytes. Median follow-up of the patients was 3.51 years (2.73; 4.48) with 34 deaths. Absolute mortality risk increases in patients displaying a MD-2 serum concentration greater than the median (302 ng/ml) was 23% (P < 0.0001). Age- and sex-adjusted Cox regression analyses demonstrated that mortality risk was highly related to MD-2 concentrations (P < 0.001), but not to age or sex. An increase of 100 ng/ml in the MD-2 level was associated with an absolute mortality risk increase of 50.4%. Receiver operating characteristic (ROC) analyses showed no difference between MD-2 and nterminal-pro brain natriuretic peptide (NT-pro-BNP), while the combination of both MD-2 and NT-pro-BNP resulted in a significantly increased capability of risk prediction when compared to NT-pro-BNP alone (P = 0.014). In-vitro, recombinant MD-2 decreases cell shortening and modulates cytokine activation in isolated cardiomyocytes. CONCLUSION: MD-2 predicts long-term outcome in DCM patients and improves mortality risk prediction capability compared to NT-pro-BNP alone. In addition, MD-2 exerts direct negative inotropic effects on isolated cardiomyocytes in-vitro. Further randomized trials should confirm MD-2 as a diagnostic and therapeutic target.

Takayasu's arteritis: a case with relapse after urgent coronary revascularization.

BACKGROUND: Vasculitides are commonly unrecognized causes of coronary stenosis and myocardial ischemia. We report on a 24-year old patient with Takayasu's arteritis who underwent urgent percutaneous coronary intervention, suffered from symptomatic restenosis of the left main coronary artery during standard immunosuppressive therapy. CASE PRESENTATION: A 24-year old woman was referred for coronary angiography because of typical progressive angina pectoris. On bicycle ergometry, there were both reproducible symptoms and deep ST segment depressions on precordial leads. Semi-selective angiography of the left coronary artery revealed high-grade ostial stenosis. Because of persistent angina pectoris and electrocardiographic signs of acute myocardial ischemia, immediate percutaneous coronary angioplasty with subsequent implantation of an everolimus-eluting stent was performed. This intervention was performed with excellent angiographic results. Because of several concomitant criteria including hypoechogenicity on postprocedural intravascular ultrasonography, the diagnosis of Takayasu's disease was made. The patient was treated with prednisolone and cyclophosphamide for 5 months. Because of recurrent angina pectoris, another coronary angiography was performed, which revealed high-grade in-stent-restenosis. Immunomodulatory therapy was switched to high-dose prednisolone and the anti-IL-6 receptor antagonist tocilizumab. The high-grade in-stent-restenosis persisted, and aortocoronary bypass graft surgery was performed with two saphenous vein grafts to the left anterior descending and circumflex artery. Since then, the patient has been doing well for 2 years. CONCLUSION: In cases of treatment refractoriness during standard immunosuppressive therapy, more recently developed biological compounds may offer an alternative strategy.

Endomyocardial proteomic signature corresponding to the response of patients with dilated cardiomyopathy to immunoadsorption therapy.

Dilated cardiomyopathy (DCM) is a disease of the myocardium with reduced left ventricular ejection fraction (LVEF). Cardiac autoantibodies (AAbs) play a causal role in the development and progression of DCM. Removal of AAbs using immunoadsorption (IA/IgG) has been shown as a therapeutic option to improve cardiac function. However, the response to therapy differs significantly among patients. The reasons for this variability are not completely understood. Hitherto, no potential biomarker is available to predict improvement of cardiac function after therapy accurately. This shotgun proteome study aims to disclose the differences in the endomyocardial proteome between patients with improved LVEF after IA/IgG (responders) and those without improvement (non-responders) before therapy start. Comparative analysis revealed 54 differentially abundant proteins that were mostly confined to carbohydrate and lipid metabolism, energy and immune regulation, and cardioprotection. Selected proteins representing various functional categories were further confirmed by multiple reaction monitoring (MRM). Among those, protein S100-A8, perilipin-4, and kininogen-1 were found the most robust candidates differentiating responders and non-responders. Receiver operating characteristic curve (ROC) analysis of these proteins revealed highest potential for protein S100-A8 (AUC 0.92) with high sensitivity and specificity to be developed as a classifier for the prediction of cardiac improvement after IA/IgG therapy. SIGNIFICANCE: We evaluated the differences in the myocardial proteome of responder and non-responder DCM patients before immunoadsorption therapy and identified a number of differentially abundant proteins involved in energy and lipid metabolism, immune system, and cardioprotection. MRM was used for verification of results. Proteins S100-A8, perilipin-4, and kininogen-1 were found to display the largest differences. The results provide a lead for further studies to screen for protein biomarker candidates in plasma that might be helpful to stratify patients for immunoadsorption therapy treatment.

Immunoadsorption therapy in dilated cardiomyopathy.

Dilated cardiomyopathy is a common myocardial disease characterized by ventricular chamber enlargement and systolic dysfunction that result in heart failure. In addition to genetic predisposition, viral infection and myocardial inflammation play a causal role in the disease process of dilated cardiomyopathy. Experimental and clinical studies suggest that activation of the humoral immune system, with production of circulating cardiac autoantibodies, plays an important functional role in the development and progression of cardiac dysfunction in patients with dilated cardiomyopathy. Small open-controlled studies showed that removal of circulating antibodies by immunoadsorption results in improvement of cardiac function and decrease in myocardial inflammation. At present, immunoadsorption is an experimental treatment option for improvement of cardiac function - therapy that calls for confirmation by a placebo-controlled multicenter study.

Non-thrombotic fatal pulmonary embolism with a 'rule-out' CT scan.

We report on a 36-year-old woman suffering from metastasized breast cancer and severe dyspnoea owing to right heart failure. Symptoms and findings were highly suggestive of pulmonary embolism. In rare cases, pulmonary embolism is caused not by migration of venous thrombi but by emboli of other origin. Patients with cancer can suffer from non-thrombotic pulmonary embolism, either by (macroscopic) embolization of tumour mass or by microembolism also known as microscopic tumour microangiopathy. In patients with cancer with clinical presentation highly suggestive of pulmonary embolism, with echocardiographic findings confirming right ventricular dysfunction, and with negative CT angiography, pulmonary tumour microembolism should be considered as possible diagnosis.