Galectin-3 levels and long-term all-cause mortality and hospitalization in heart failure patients: a meta-analysis.

AIMS: This meta-analysis investigated the dose-response relationship between circulating galectin-3 levels and adverse outcomes in patients with heart failure (HF). METHODS AND RESULTS: PubMed and Embase were screened for studies on galectin-3 and HF. The outcomes of interest were all-cause mortality (ACM), and all-cause mortality or HF-related rehospitalization (ACM/HFR), with a follow-up time of more than 6 months. For categorical variables, comparisons between groups with the highest and lowest galectin-3 levels were pooled. For continuous variables, the risks of ACM and ACM/HFR increase per 1-standard deviation (SD) and 1-unit after logarithmic transformation galectin-3 levels were pooled. A random-effects model was employed to calculate the pooled results, and all pooled results were expressed as hazard ratios (HRs) and 95% confidence intervals (CIs). Besides, a dose-response analysis was performed. Twenty-four cohort studies were included. In HF patients, higher circulating galectin-3 levels were significantly associated with a higher risk of long-term ACM (HR, 1.65; 95% CI 1.28-2.13; I2 = 66%), and 1 ng/mL increase in galectin-3 was associated with a 4% (HR, 1.04; 95% CI 1.02-1.06; P = 0.002) increase in hazard. Similarly, higher circulating galectin-3 levels were significantly associated with a higher risk of long-term ACM/HFR (HR, 1.52; 95% CI, 1.15 to 2.00; I2 = 76%), and 1 ng/mL increase in galectin-3 was associated with a 3% (HR, 1.03; 95% CI 1.02-1.04; P < 0.001) increase in hazard. An increase of 1-SD in galectin-3 units was associated with a 29% increased hazard of long-term ACM (HR 1.29; 95% CI 1.13-1.48; I2 = 42%) and a 22% increased hazard of ACM/HFR (HR 1.22; 95% CI 1.07-1.38; I2 = 60%). Similarly, an increase of 1-log in galectin-3 units was associated with a 98% higher hazard of long-term ACM (HR 1.98; 95% CI 1.48-2.65; I2 = 41%) and an 83% higher hazard of ACM/HFR in HF patients (HR 1.83; 95% CI 1.02-3.28; I2 = 7%). Correlation analysis showed a moderate positive correlation between baseline galectin-3 and N terminal pro brain natriuretic peptide levels (r = 0.48, P = 0.045) and a weak negative correlation with eGFR (r = -0.39, P = 0.077). CONCLUSIONS: Higher circulating galectin-3 levels after hospitalization of HF patients are linearly and positively associated with the risk of long-term ACM and ACM/HFR.

Amphetamine increases vascular permeability by modulating endothelial actin cytoskeleton and NO synthase via PAR-1 and VEGF-R.

Abuse of amphetamine-type stimulants is linked to cardiovascular adverse effects like arrhythmias, accelerated atherosclerosis, acute coronary syndromes and sudden cardiac death. Excessive catecholamine release following amphetamine use causes vasoconstriction and vasospasms, over time leading to hypertension, endothelial dysfunction or even cardiotoxicity. However, immediate vascular pathomechanisms related to amphetamine exposure, especially endothelial function, remain incompletely understood and were analyzed in this study. Pharmaco-pathological effects of acute d-amphetamine-sulfate (DAM) were investigated ex vivo using contraction-force measurements of rat carotid artery rings and in vitro using label-free, real-time electrochemical impedance spectroscopy (EIS) on endothelial and smooth muscle cells. Specific receptor and target blocking was used to identify molecular targets and to characterize intracellular signaling. DAM induced vasodilation represented by 29.3±2.5% decrease in vascular tone (p<0.001) involving vascular endothelial growth factor receptor (VEGF-R) and protease activated receptor 1 (PAR-1). EIS revealed that DAM induces endothelial barrier disruption (-75.9±1.1% of initial cellular impedance, p<0.001) also involving VEGF-R and PAR-1. Further, in response to DAM, Rho-associated protein kinase (ROCK) mediated reversible contraction of actin cytoskeleton resulting in endothelial barrier disruption. Dephosphorylation of Serine1177 (-50.8±3.7%, p<0.001) and Threonine495 (-44.8±6.5%, p=0.0103) of the endothelial NO synthase (eNOS) were also observed. Blocking of VEGF-R and PAR-1 restored baseline eNOS Threonine495 phosphorylation. DAM induced vasodilation, enhanced vascular permeability and actin cytoskeleton contraction and induced eNOS hypophosphorylation involving VEGF-R, PAR-1 and ROCK. These results may contribute to a better understanding of severe adverse cardiovascular effects in amphetamine abuse.

Impact of anaemia and iron deficiency on outcomes in cardiogenic shock complicating acute myocardial infarction.

AIMS: Anaemia and iron deficiency (ID) are common comorbidities in cardiovascular patients and are associated with a poor clinical status, as well as a worse outcome in patients with heart failure and acute myocardial infarction (AMI). Nevertheless, data concerning the impact of anaemia and ID on clinical outcomes in patients with cardiogenic shock (CS) are scarce. This study aimed to assess the impact of anaemia and ID on clinical outcomes in patients with CS complicating AMI. METHODS AND RESULTS: The presence of anaemia (haemoglobin <13 g/dl in men and <12 g/dl in women) or ID (ferritin <100 ng/ml or transferrin saturation <20%) was determined in patients with CS due to AMI from the CULPRIT-SHOCK trial. Blood samples were collected in the catheterization laboratory during initial percutaneous coronary intervention. Clinical outcomes were compared in four groups of patients having neither anaemia nor ID, against patients with anaemia with or without ID and patients with ID only. A total of 427 CS patients were included in this analysis. Anaemia without ID was diagnosed in 93 (21.7%), anaemia with ID in 54 study participants (12.6%), ID without anaemia in 72 patients (16.8%), whereas in 208 patients neither anaemia nor ID was present (48.9%). CS patients with anaemia without ID were older (73 ± 10 years, p = 0.001), had more frequently a history of arterial hypertension (72.8%, p = 0.01), diabetes mellitus (47.8%, p = 0.001), as well as chronic kidney disease (14.1%, p = 0.004) compared to CS patients in other groups. Anaemic CS patients without ID presence were at higher risk to develop a composite from all-cause death or renal replacement therapy at 30-day follow-up (odds ratio [OR] 3.83, 95% confidence interval [CI] 2.23-6.62, p < 0.001) than CS patients without anaemia/ID. The presence of ID in CS patients, with and without concomitant anaemia, did not increase the risk for the primary outcome (OR 1.17, 95% CI 0.64-2.13, p = 0.64; and OR 1.01, 95% CI 0.59-1.73, p = 0.54; respectively) within 30 days of follow-up. In time-to-event Kaplan-Meier analysis, anaemic CS patients without ID had a significantly higher hazard ratio (HR) for the primary outcome (HR 2.11, 95% CI 1.52-2.89, p < 0.001), as well as for death from any cause (HR 1.90, 95% CI 1.36-2.65, p < 0.001) and renal replacement therapy during 30-day follow-up (HR 2.99, 95% CI 1.69-5.31, p < 0.001). CONCLUSION: Concomitant anaemia without ID presence in patients with CS at hospital presentation is associated with higher risk for death from any cause or renal replacement therapy and the individual components of this composite endpoint within 30 days after hospitalization. ID has no relevant impact on clinical outcomes in patients with CS.

Nuclear magnetic resonance spectroscopy to quantify major extracellular matrix components in fibro-calcific aortic valve disease.

Fibro-calcific aortic valve disease (FCAVD) is a pathological condition marked by overt fibrous and calcific extracellular matrix (ECM) accumulation that leads to valvular dysfunction and left ventricular outflow obstruction. Costly valve implantation is the only approved therapy. Multiple pharmacological interventions are under clinical investigation, however, none has proven clinically beneficial. This failure of translational approaches indicates incomplete understanding of the underlying pathomechanisms and may result from a limited toolbox of scientific methods to assess the cornerstones of FCAVD: lipid deposition, fibrous and calcific ECM accumulation. In this study, we evaluated magic-angle spinning (MAS) nuclear magnetic resonance (NMR) spectroscopy to both, qualitatively and quantitatively assess these key elements of FCAVD pathogenesis. NMR spectra showed collagen, elastin, triacylglycerols, and phospholipids in human control and FCAVD tissue samples (n = 5). Calcification, measured by the hydroxyapatite content, was detectable in FCAVD tissues and in valve interstitial cells under procalcifying media conditions. Hydroxyapatite was significantly higher in FCAVD tissues than in controls (p < 0.05) as measured by 31P MAS NMR. The relative collagen content was lower in FCAVD tissues vs. controls (p < 0.05). Overall, we demonstrate the versatility of NMR spectroscopy as a diagnostic tool in preclinical FCAVD assessment.

C-kitpos cells in the human left atrial appendage.

Background: Subpopulations of myocardial c-kitpos cells have the ability to stimulate regeneration in ischemic heart disease by paracrine effects. The left atrial appendage (LAA), which is easy accessible during cardiac surgery, may represent a perfect source for c-kitpos cell extraction for autologous cell therapies in the living human. So far, frequency and distribution of c-kitpos cells in LAA are unknown. Methods: LAAs of patients who underwent cardiac surgery due to coronary artery disease (coronary artery bypass graft, CABG), valvular heart disease or both and of two body donors were examined. Tissue was fixed in 4 % paraformaldehyde, embedded in paraffin, dissected in consecutive sections and stained for c-kitpos cells. In parallel, grade of fibrosis, amount of fat per section and cells positive for mast cell tryptase were examined. Results: We collected 27 LAAs (37.0 % female, mean left ventricular ejection fraction 50.4 %, 63.0 % persistent atrial fibrillation (AF)). Most of the patients underwent combined CABG and valve surgery (51.9 %). C-kitpos cells were detected in 3 different regions: A) Attached to the epicardial fat layer, B) close to vascular structures and C) between cardiomyocytes. C-kitpos cells ranged from 0.05 c-kitpos cells per mm2 to 67.5 c-kitpos cells per mm2. We found no association between number of c-kitpos cells and type of AF, amount of fibrosis or amount of fat. Up to 72 % of c-kitpos cells also showed a positive staining for mast cell tryptase. Conclusion: C-kitpos cells are frequent in LAAs of cardiovascular patients with a rather homogenous distribution throughout the LAA. The LAA can therefore be considered as a source for extraction of a reasonable quantity of autologous cardiac progenitor cells in the living human patient.

Systemic Effects of Homoarginine Supplementation on Arginine Metabolizing Enzymes in Rats with Heart Failure with Preserved Ejection Fraction.

A restoration of low homoarginine (hArg) levels in obese ZSF1 rats (O-ZSF1) before (S1-ZSF1) and after (S2-ZSF1) the manifestation of heart failure with preserved ejection fraction (HFpEF) did not affect the worsening of cardiac HFpEF characteristics. Here, potential regulation of key enzymes of arginine metabolism in other organs was analyzed. Arginase 2 (ARG2) was reduced >35% in the kidney and small intestine of hArg-supplemented rats compared to O-ZSF1. Glycine amidinotransferase (GATM) was 29% upregulated in the kidneys of S1-ZSF1. Dimethylarginine dimethylaminohydrolase 1 (DDAH1) levels were reduced >50% in the livers of O-ZSF1 but restored in S2-ZSF1 compared to healthy rats (L-ZSF1). In the skeletal muscle, iNOS was lower in O-ZSF1 and further decreased in S1-ZSF1 and S2-ZSF1 compared to L-ZSF1. iNOS levels were lower in the liver of the S2-ZSF1 group but higher in the kidneys of S1-ZSF1 compared to L-ZSF1. Supplementation with hArg in an in vivo HFpEF model resulted in the inhibition of renal ARG2 and an increase in GATM expression. This supplementation might contribute to the stabilization of intestinal iNOS and ARG2 imbalances, thereby enhancing barrier function. Additionally, it may offer protective effects in skeletal muscle by downregulating iNOS. In the conceptualization of hArg supplementation studies, the current disease progression stage as well as organ-specific enzyme regulation should be considered.

High resolution monitoring of valvular interstitial cell driven pathomechanisms in procalcific environment using label-free impedance spectroscopy.

Introduction: Fibro-calcific aortic valve disease has high prevalence and is associated with significant mortality. Fibrotic extracellular matrix (ECM) remodeling and calcific mineral deposition change the valvular microarchitecture and deteriorate valvular function. Valvular interstitial cells (VICs) in profibrotic or procalcifying environment are frequently used in vitro models. However, remodeling processes take several days to weeks to develop, even in vitro. Continuous monitoring by real-time impedance spectroscopy (EIS) may reveal new insights into this process. Methods: VIC-driven ECM remodeling stimulated by procalcifying (PM) or profibrotic medium (FM) was monitored by label-free EIS. Collagen secretion, matrix mineralization, viability, mitochondrial damage, myofibroblastic gene expression and cytoskeletal alterations were analyzed. Results and Discussion: EIS profiles of VICs in control medium (CM) and FM were comparable. PM reproducibly induced a specific, biphasic EIS profile. Phase 1 showed an initial impedance drop, which moderately correlated with decreasing collagen secretion (r = 0.67, p = 0.22), accompanied by mitochondrial membrane hyperpolarization and cell death. Phase 2 EIS signal increase was positively correlated with augmented ECM mineralization (r = 0.97, p = 0.008). VICs in PM decreased myofibroblastic gene expression (p < 0.001) and stress fiber assembly compared to CM. EIS revealed sex-specific differences. Male VICs showed higher proliferation and in PM EIS decrease in phase 1 was significantly pronounced compared to female VICs (male minimum: 7.4 ± 4.2%, female minimum: 26.5 ± 4.4%, p < 0.01). VICs in PM reproduced disease characteristics in vitro remarkably fast with significant impact of donor sex. PM suppressed myofibroblastogenesis and favored ECM mineralization. In summary, EIS represents an efficient, easy-to-use, high-content screening tool enabling patient-specific, subgroup- and temporal resolution.

Clonal Hematopoiesis Mutations Are Present in Atherosclerotic Lesions in Peripheral Artery Disease.

Clonal hematopoiesis (CH)-associated mutations increase the risk of atherosclerotic cardiovascular diseases. However, it is unclear whether the mutations detected in circulating blood cells can also be detected in tissues associated with atherosclerosis, where they could affect physiology locally. To address this, the presence of CH mutations in peripheral blood, atherosclerotic lesions and associated tissues was assessed in a pilot study of 31 consecutive patients with peripheral vascular disease (PAD) who underwent open surgical procedures. Next-generation sequencing was used to screen the most commonly mutated loci (DNMT3A, TET2, ASXL1 and JAK2). Twenty CH mutations were detected in peripheral blood of 14 (45%) patients, 5 of whom had more than one mutation. TET2 (11 mutations, 55%) and DNMT3A (8 mutations, 40%) were the most frequently affected genes. Altogether, 88% of the mutations detectable in peripheral blood were also present in the atherosclerotic lesions. Twelve patients also had mutations in perivascular fat or subcutaneous tissue. The presence of CH mutations in PAD-associated tissues as well as in blood suggests that CH mutations may make a previously unknown contribution to PAD disease biology.

Circulating Galectin-3 in Patients with Cardiogenic Shock Complicating Acute Myocardial Infarction Treated with Mild Hypothermia: A Biomarker Sub-Study of the SHOCK-COOL Trial.

Background: Galectin-3 (Gal-3) is considered a potential cardiovascular inflammatory marker that may provide additional risk stratification for patients with acute heart failure. It is unknown whether mild therapeutic hypothermia (MTH) impacts Gal-3 levels. Therefore, this biomarker study aimed to investigate the effect of MTH on Gal-3. Methods: In the randomized SHOCK-COOL trial, 40 patients with cardiogenic shock (CS) complicating acute myocardial infraction (AMI) were randomly assigned to the MTH (33 °C) or control group in a 1:1 ratio. Blood samples were collected on the day of admission/day 1, day 2, and day 3. Gal-3 level kinetics throughout these time points were compared between the MTH and control groups. Additionally, potential correlations between Gal-3 and clinical patient characteristics were assessed. Multiple imputations were performed to account for missing data. Results: In the control group, Gal-3 levels were significantly lower on day 3 than on day 1 (day 1 vs. day 3: 3.84 [IQR 2.04−13.3] vs. 1.79 [IQR 1.23−3.50] ng/mL; p = 0.049). Gal-3 levels were not significantly different on any day between the MTH and control groups (p for interaction = 0.242). Spearman's rank correlation test showed no significant correlation between Gal-3 levels and sex, age, smoking, body mass index (BMI), and levels of creatine kinase-MB, creatine kinase, C-reactive protein, creatinine, and white blood cell counts (all p > 0.05). Patients with lower Gal-3 levels on the first day after admission demonstrated a higher risk of all-cause mortality at 30 days (hazard ratio, 2.67; 95% CI, 1.11−6.42; p = 0.029). In addition, Gal-3 levels on day 1 had a good predictive value for 30-day all-cause mortality with an area under the receiver operating characteristic curve of 0.696 (95% CI: 0.513−0.879), with an optimal cut-off point of less than 3651 pg/mL. Conclusions: MTH has no effect on Gal-3 levels in patients with CS complicating AMI compared to the control group. In addition, Gal-3 is a relatively stable biomarker, independent of age, sex, and BMI, and Gal-3 levels at admission might predict the risk of 30-day all-cause mortality.

Impact of Clonal Hematopoiesis in Patients With Cardiogenic Shock Complicating Acute Myocardial Infarction.

BACKGROUND: Clonal hematopoiesis of indeterminate potential (CHIP) is common in elderly individuals and is associated with an increased risk of both hematologic malignancies and cardiovascular disease. The impact of CHIP on the outcomes for patients with cardiogenic shock (CS) complicating acute myocardial infarction (AMI) remains undetermined. OBJECTIVES: The purpose of this study was to determine the prognostic impact of CHIP in CS after AMI. METHODS: Blood samples were obtained at randomization from 446 patients included in the CULPRIT-SHOCK (Culprit Lesion Only vs Multivessel Percutaneous Coronary Intervention in Cardiogenic Shock; NCT01927549) trial. CHIP was assessed using a next-generation sequencing approach targeting the most commonly mutated genes; the primary outcome at 30 days comprised all-cause mortality and renal replacement therapy. RESULTS: CHIP variants at ≥2% variant allele frequency were detected in 29% (n = 129), most commonly in the DNMT3A or TET2 genes, which harbored 47% and 36% of all mutations, respectively. Compared to non-CHIP patients, CHIP carriers were older and had decreased renal function and increased levels of N-terminal pro-B-type natriuretic peptide and inflammatory biomarkers. CHIP carriers had worse short-term outcomes measured either as mortality or as the combined clinical endpoint of mortality or severe renal failure within 30 days. Association of CHIP with the combined endpoint was independent of age and biomarkers reflecting kidney function, heart failure severity, and inflammation (OR: 1.83; 95% CI: 1.05-3.21; P = 0.03) but not significant regarding all-cause mortality (OR: 1.67; 95% CI: 0.96-2.90; P = 0.069). CONCLUSIONS: CHIP is frequent among AMI and CS patients and is associated with impaired clinical outcome. CHIP assessment may facilitate risk stratification in patients with CS and imply novel treatment targets. (Culprit Lesion Only vs Multivessel Percutaneous Coronary Intervention in Cardiogenic Shock [CULPRIT-SHOCK]; NCT01927549).

Tubulin expression and modification in heart failure with preserved ejection fraction (HFpEF).

Diastolic dysfunction in heart failure with preserved ejection fraction (HFpEF) is characterised by increased left ventricular stiffness and impaired active relaxation. Underpinning pathomechanisms are incompletely understood. Cardiac hypertrophy and end stage heart disease are associated with alterations in the cardiac microtubule (MT) network. Increased amounts and modifications of α-tubulin associate with myocardial stiffness. MT alterations in HFpEF have not been analysed yet. Using ZSF1 obese rats (O-ZSF1), a validated HFpEF model, we characterised MT-modifying enzymes, quantity and tyrosination/detyrosination pattern of α-tubulin at 20 and 32 weeks of age. In the left ventricle of O-ZSF1, α-tubulin concentration (20 weeks: 1.5-fold, p = 0.019; 32 weeks: 1.7-fold, p = 0.042) and detyrosination levels (20 weeks: 1.4-fold, p = 0.013; 32 weeks: 1.3-fold, p = 0.074) were increased compared to lean ZSF1 rats. Tyrosination/α-tubulin ratio was lower in O-ZSF1 (20 weeks: 0.8-fold, p = 0.020; 32 weeks: 0.7-fold, p = 0.052). Expression of α-tubulin modifying enzymes was comparable. These results reveal new alterations in the left ventricle in HFpEF that are detectable during early (20 weeks) and late (32 weeks) progression. We suppose that these alterations contribute to diastolic dysfunction in HFpEF and that reestablishment of MT homeostasis might represent a new target for pharmacological interventions.

Effects of homoarginine supplementation on heart and skeletal muscle of rats with heart failure with preserved ejection fraction.

AIM: Heart failure with preserved ejection fraction (HFpEF) is associated with left ventricular stiffness, impaired diastolic relaxation, and severe exercise intolerance. Decreased homoarginine (hArg) levels are an independent predictor of mortality in cardiovascular disease and correlate with impaired exercise performance. We recently reported alterations in arginine, hArg, and related amino acids in obese ZSF1 rats (O-ZSF1), with a HFpEF phenotype. Although low hArg is associated with diastolic dysfunction in humans, potential effects of hArg supplementation were not tested yet. METHODS AND RESULTS: At an age of 6 weeks, 12 O-ZSF1 were randomized into two groups: (1) O-ZSF1 rats supplemented with hArg in their drinking water (sO-ZSF1) or (2) O-ZSF1 rats receiving no hArg supplementation (O-ZSF1). At an age of 32 weeks, effects of primary prevention by hArg supplementation on echocardiographic, histological, and functional parameters of heart and skeletal muscle were determined. Lean ZSF1 rats (L-ZSF1) served as controls. hArg supplementation did not prevent impairment of diastolic relaxation (E/e': O-ZSF1 21 ± 3 vs. sO-ZSF1 22 ± 3, P = 0.954, L-ZSF1 18 ± 5) but resulted in more cardiac fibrosis (histological collagen staining: +57% in sO-ZSF1 vs. O-ZSF1, P = 0.027) and increased collagen gene expression (Col1a1: +48% in sO-ZSF1 vs. O-ZSF1, P = 0.026). In contrary, right ventricular function was preserved by hArg supplementation (TAPSE (mm): O-ZSF1 1.2 ± 0.3 vs. sO-ZSF1 1.7 ± 0.3, P = 0.020, L-ZSF1 1.8 ± 0.4). Musculus soleus maximal specific muscle force (N/cm2 ) in O-ZSF1 (30.4 ± 0.8) and sO-ZSF1 (31.9 ± 0.9) was comparable but significantly reduced compared with L-ZSF1 (36.4 ± 0.7; both P < 0.05). Maximal absolute muscle force (g) (O-ZSF1: 177.6 ± 7.8, sO-ZSF1: 187.8 ± 5.0, L-ZSF1: 181.5 ± 7.9, all P > 0.05) and cross-sectional fibre area (arbitrary units) (O-ZSF1: 1697 ± 57, sO-ZSF1: 1965 ± 121, L-ZSF1: 1691 ± 104, all P > 0.05) were not altered. CONCLUSIONS: Preservation of physiological hArg level in HFpEF may not be suited to prevent alterations in left ventricular and skeletal muscle function and structure. However, hArg supplementation may be beneficial for right ventricular function especially in pulmonary hypertension in HFpEF. We may speculate that clinically observed decreased hArg level are not the cause but the consequence of a yet unrecognized pathomechanism that underpins HFpEF.

Soluble ST2 Receptor: Biomarker of Left Ventricular Impairment and Functional Status in Patients with Inflammatory Cardiomyopathy.

INTRODUCTION: Inflammatory cardiomyopathy (ICM) frequently leads to myocardial fibrosis, resulting in permanent deterioration of the left ventricular function and an unfavorable outcome. Soluble suppression of tumorigenicity 2 receptor (sST2) is a novel marker of inflammation and fibrosis in cardiovascular tissues. sST2 was found to be helpful in predicting adverse outcomes in heart failure patients with reduced ejection fraction. The aim of this study was to determine the association of sST2 plasma levels with cardiac magnetic resonance (CMR) and echocardiography imaging features of left ventricular impairment in ICM patients, as well as to evaluate the applicability of sST2 as a prognosticator of the clinical status in patients suffering from ICM. METHODS: We used plasma samples of 89 patients presenting to the Heart Center Leipzig with clinically suspected myocardial inflammation. According to immunohistochemical findings in endomyocardial biopsies (EMB) conducted in the context of patients' diagnostic work-up, inflammatory cardiomyopathy was diagnosed in 60 patients (ICM group), and dilated cardiomyopathy in 29 patients (DCM group). All patients underwent cardiac catheterization for exclusion of coronary artery disease and CMR imaging on 1.5 or 3 Tesla. sST2 plasma concentration was determined using ELISA. RESULTS: Mean plasma concentration of sST2 in the whole patient cohort was 45.8 ± 26.4 ng/mL (IQR 27.5 ng/mL). In both study groups, patients within the highest quartile of sST2 plasma concentration had a significantly lower left ventricular ejection fraction (LV-EF) compared to patients within the lowest sST2 plasma concentration quartile (26 ± 11% vs. 40 ± 13%, p = 0.05 for ICM and 24 ± 13% vs. 51 ± 10%, p = 0.004 for DCM). sST2 predicted New York Heart Association (NYHA) class III/IV at 12 months follow-up more efficiently in ICM compared to DCM patients (AUC 0.85 vs. 0.61, p = 0.02) and was in these terms superior to NT-proBNP and cardiac troponin T. ICM patients with sST2 plasma concentration higher than 44 ng/mL at baseline had a significantly higher probability of being assigned to NYHA class III/IV at 12 months follow-up (hazard ratio 2.8, 95% confidence interval 1.01-7.6, log rank p = 0.05). CONCLUSION: Plasma sST2 levels in ICM patients reflect the degree of LV functional impairment at hospital admission and predict functional NYHA class at mid-term follow-up. Hence, ST2 may be helpful in the evaluation of disease severity and in the prediction of the clinical status in ICM patients.

Anti-inflammatory HDL effects are impaired in atrial fibrillation.

High-density lipoprotein (HDL), best known for cholesterol transport, also has anti-inflammatory effects. Previous studies suggest involvement of myeloperoxidase (MPO) in modification of HDL. HDL bound Sphingosine-1-phosphate (S1P) has been implied to be an essential protein regarding beneficial HDL effects. In this study, we analyzed anti-inflammatory HDL properties in patients with atrial fibrillation (AF), a disease involving atrial inflammation, compared to non-AF controls and whether anti-inflammatory properties improve upon catheter ablation. Additionally, association with serum concentrations of MPO and S1P were assessed. We isolated HDL from 25 AF patients, 13 non-AF individuals and 14 AF patients at follow-up (FU) after catheter ablation. S1P was measured in a cohort of 141 AF and 21 FU patients. Following preincubation with HDL from either group, bovine aortic endothelial cells were stimulated using tumor necrosis factor α and expression of pro-inflammatory genes intercellular adhesion molecule 1 (ICAM1), vascular cell adhesion molecule 1 (VCAM1), E-selectin (SELE) and P-selectin (SELP) was assessed using qPCR. Concentrations of circulating protein of these genes as well as MPO and S1P were measured in serum samples. Compared to non-AF individuals HDL from AF patients suppressed gene expression of the pro-inflammatory adhesion molecules ICAM1, VCAM1, SELE and SELP 27%, 18%, 21% and 57% less, respectively (p < 0.05 for all except SELE p = 0.06). In FU patients, the anti-inflammatory HDL activity was improved (suppression of ICAM1 + 22%, VCAM1 + 10%, SELE + 38% and SELP + 75%, p < 0.05 for all except VCAM1 p = 0.08). AF patients using angiotensin converting enzyme inhibitors or angiotensin receptor blockers had better anti-inflammatory HDL properties than non-users (gene expression suppression at least 28% more, p < 0.05 for all except ICAM1 p = 0.051). Circulating protein concentrations were not correlated with in vitro gene-expression, but circulating P-selectin was generally elevated in AF and FU patients compared to non-AF patients. MPO plasma concentration was positively associated with gene-expression of ICAM1, VCAM1 and SELP (r2 > 0.4, p < 0.05). Serum concentrations of S1P were increased in FU patients {1.201 µM [1.077-1.543]} compared to AF patients {0.953 µM [0.807-1.135], p < 0.01} but not correlated with ICAM1, VCAM1 and SELP gene expression. We conclude that the anti-inflammatory activity of HDL is impaired in AF patients, which might promote AF progression and AF-associated complications.

NT-proANP levels in peripheral and cardiac circulation.

BACKGROUND: Recent studies have reported an association between N-terminal atrial natriuretic peptide (NT-proANP) and the progression of atrial fibrillation (AF). However, NT-proANP levels in peripheral and cardiac circulation in AF patients and in non-AF individuals need to be defined. The aims of the current study are (1) to analyze NT-proANP levels in peripheral and cardiac circulation in AF patients and (2) to compare NT-proANP levels in individuals with and without AF. METHODS: We recruited AF patients who were undergoing their first AF catheter ablation and non-AF individuals. Blood plasma samples taken from the femoral vein and the left atrium (LA) were collected before AF ablation in the AF patients and from the cubital vein in the non-AF controls. Low voltage areas (LVAs) were determined using high-density maps during catheter ablation and defined as < 0.5 mV. RESULTS: The study included 189 AF patients (64 ± 10 years, 59% male, 61% persistent AF, 30% LVAs) and 26 non-AF individuals (58 ± 10 years, 50% male). Patients with AF were significantly older and had larger LA (p < 0.05). Compared to non-AF controls, peripheral and cardiac NT-proANP levels were significantly higher in AF patients without and with LVAs (median 5.4, 10.5, 14.8 ng/ml, respectively, p < 0.001). In multivariable analysis, NT-proANP (OR 1.238, 95% CI 1.007-1.521, p = 0.043) remained significantly different between non-AF individuals and AF patients. In AF, NT-proANP levels were significantly higher in the cardiac blood samples than in the peripheral blood (median 13.0 versus 11.4 ng/ml, p = 0.003). The ability to predict LVAs was modest when using cardiac NT-proANP (AUC 0.661) and peripheral NT-proANP (AUC 0.635), without statistical difference (p = 0.937). CONCLUSIONS: NT-proANP levels are higher in individuals with AF than in controls and are more pronounced in progressed AF. Elevated cardiac and peripheral NT-proANP levels similarly predict LVAs.

Profile of urinary amino acids and their post-translational modifications (PTM) including advanced glycation end-products (AGEs) of lysine, arginine and cysteine in lean and obese ZSF1 rats.

Heart failure with preserved ejection fraction (HFpEF) is associated with high mortality and has an increasing prevalence associated with the demographic change and limited therapeutic options. Underlying mechanisms are largely elusive and need to be explored to identify specific biomarkers and new targets, which mirror disease progression and intervention success. Obese ZSF1 (O-ZSF1) rats are a useful animal model, as they spontaneously develop hypertension, hyperlipidemia and glucose intolerance and finally HFpEF. The urinary profile of amino acids and their metabolites of post-translational modifications (PTM), including the advanced glycation end-products (AGEs) of lysine, arginine and cysteine, are poorly investigated in HFpEF and ZSF1 rats. The aim of the present study was to characterize the status of free amino acids and their metabolites of PTM and glycation in lean ZSF1 (L-ZSF1) and O-ZSF1 rats in urine aiming to find possible effects of glucose on the excretion of native and modified amino acids. In the urine of twelve L-ZSF1 and twelve O-ZFS1 rats collected at the age of 20 weeks, we measured the concentration of native and modified amino acids by reliable previously validated stable-isotope dilution gas chromatography-mass spectrometry (GC-MS) approaches. Serum glucose was 1.39-fold higher in the O-ZSF1 rats, while urinary creatinine concentration was 2.5-fold lower in the O-ZSF1 rats. We observed many differences in urinary amino acids excretion between L-ZSF1 and O-ZSF1 rats. The creatinine-corrected homoarginine excretion was twofold lower in the O-ZSF1 rats. We also observed distinct associations between the concentrations of serum glucose and urinary amino acids including their PTM and AGE metabolites in the L-ZSF1 and O-ZSF1 rats. Our study shows that PTM metabolites and AGEs are consistently lower in the L-ZSF1 than in the O-ZSF1 rats. Serum malondialdehyde (MDA) concentration was higher in the O-ZSF1 rats. These results suggest that hyperglycemia, hyperlipidemia and elevated oxidative stress in the O-ZSF1 rats favor PTM methylation of arginine and lysine and the glycation of lysine and cysteine. The area under the receiver operation characteristic (ROC) curve values were 0.996 for serum glucose, 0.951 for urinary creatinine, 0.939 for serum MDA, 0.885 for Nε-carboxyethyl-lysine, 0.830 for carboxyethyl-cysteine, and 0.792 for monomethyl-lysine. Non-invasive measurement of methylation and glycation products of arginine, lysine and cysteine residues in proteins in urine of L-ZSF1 and O-ZSF1 rats may be useful in studying pathophysiology and pharmacology of HFpEF.

Everolimus-Induced Immune Effects after Heart Transplantation: A Possible Tool for Clinicians to Monitor Patients at Risk for Transplant Rejection.

BACKGROUND: Patients treated with an inhibitor of the mechanistic target of rapamycin (mTORI) in a calcineurin inhibitor (CNI)-free immunosuppressive regimen after heart transplantation (HTx) show a higher risk for transplant rejection. We developed an immunological monitoring tool that may improve the identification of mTORI-treated patients at risk for rejection. METHODS: Circulating dendritic cells (DCs) and regulatory T cells (Tregs) were analysed in 19 mTORI- and 20 CNI-treated HTx patients by flow cytometry. Principal component and cluster analysis were used to identify patients at risk for transplant rejection. RESULTS: The percentages of total Tregs (p = 0.02) and CD39+ Tregs (p = 0.05) were higher in mTORI-treated patients than in CNI-treated patients. The principal component analysis revealed that BDCA1+, BDCA2+ and BDCA4+ DCs as well as total Tregs could distinguish between non-rejecting and rejecting mTORI-treated patients. Most mTORI-treated rejectors showed higher levels of BDCA2+ and BDCA4+ plasmacytoid DCs and lower levels of BDCA1+ myeloid DCs and Tregs than mTORI non-rejectors. CONCLUSION: An mTORI-based immunosuppressive regimen induced a sufficient, tolerance-promoting reaction in Tregs, but an insufficient, adverse effect in DCs. On the basis of patient-specific immunological profiles, we established a flow cytometry-based monitoring tool that may be helpful in identifying patients at risk for rejection.

Arginine metabolism and nitric oxide turnover in the ZSF1 animal model for heart failure with preserved ejection fraction.

Endothelial dysfunction and altered nitric oxide (NO) metabolism are considered causal factors in heart failure with preserved ejection fraction (HFpEF). NO synthase activity depends on the availability of arginine and its derivatives. Thus, we analyzed arginine, associated metabolites, arginine-metabolizing enzymes and NO turnover in 20-week-old female healthy lean (L-ZSF1) and obese ZSF1 rats (O-ZSF1) with HFpEF. Serum, urine and lysates of liver, kidney and heart were analyzed. There were significantly lower lysine (- 28%), arginine (- 31%), homoarginine (- 72%) and nitrite (- 32%) levels in serum of O-ZSF1 rats. Ornithine (+ 60%) and citrulline (+ 20%) levels were higher. Similar results were found in the heart. Expression of arginine consuming enzymes in liver and kidney was unchanged. Instead, we observed a 5.8-fold higher arginase 1 expression, presumably of granulocyte origin, in serum and > fourfold increased cardiac macrophage invasion in O-ZSF1. We conclude that inflammatory cells in blood and heart consume arginine and probably homoarginine via arginase 1 and inducible NO synthase and release ornithine and citrulline. In combination with evidence for decreased NO turnover in O-ZSF1 rats, we assume lower arginine bioavailability to endothelial NO synthase.