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Background: This study analyzed the utility of soluble ST2 (sST2) and GDF-15 as biomarkers of right ventricular (RV) function in patients with pulmonary hypertension (PH). Methods: GDF-15 and sST2 serum concentrations were measured in patients with PH (n = 628), dilated cardiomyopathy (n = 31) and left ventricular hypertrophy (n = 47), and in healthy controls (n = 61). Results: Median sST2 and GDF-15 levels in patients with left ventricular hypertrophy were higher than in patients with PH and dilated cardiomyopathy. In tertile analysis GDF-15 >1363 pg/ml and sST2 >38 ng/ml were associated with higher N-terminal pro-brain natriuretic peptide, RV systolic dysfunction, RV-pulmonary arterial uncoupling and hemodynamic impairment. Conclusion: GDF-15 and sST2 are potential biomarkers of RV dysfunction in patients with PH.

Background Fibroblast growth factor 23 (FGF-23) has been associated with left ventricular hypertrophy (LVH) and heart failure. However, its role in right ventricular (RV) remodeling and RV failure is unknown. This study analyzed the utility of FGF-23 as a biomarker of RV function in patients with pulmonary hypertension (PH). Methods In this observational study, FGF-23 was measured in the plasma of patients with PH (n = 627), dilated cardiomyopathy (DCM, n = 59), or LVH with severe aortic stenosis (n = 35). Participants without LV or RV abnormalities served as controls (n = 36). Results Median FGF-23 plasma levels were higher in PH patients than in healthy controls (p p 2. Association of FGF-23 with parameters of RV function was independent of the glomerular filtration rate in regression analysis. Conclusion FGF-23 may serve as a biomarker for maladaptive RV remodeling in patients with PH. Graphic abstract

Purpose Physical activity is associated with altered levels of circulating microRNAs (ci-miRNAs). Changes in miRNA expression have great potential to modulate biological pathways of skeletal muscle hypertrophy and metabolism. This study was designed to determine whether the profile of ci-miRNAs is altered after different approaches of endurance exercise. Methods Eighteen healthy volunteers (aged 24 ± 3 years) participated this three-arm, randomized-balanced crossover study. Each arm was a single bout of treadmill-based acute endurance exercise at (1) 100% of the individual anaerobic threshold (IANS), (2) at 80% of the IANS and (3) at 80% of the IANS with blood flow restriction (BFR). Load-associated outcomes (fatigue, feeling, heart rate, and exhaustion) as well as acute effects (circulating miRNA patterns and lactate) were determined. Results All training interventions increased the lactate concentration (LC) and heart rate (HR) (p p p = 0.037 and p = 0.003). The level of miR-142-5p and miR-197-3p were up-regulated in both interventions without BFR (p p = 0.038). In LI-BFR, the level of miR-342-3p and miR-424-5p was confirmed to be up-regulated (p p = 0.011, r = − 0.343/miR-199a-3p, p = 0.045, r = − 0.274/miR-125b-5p, p = 0.026, r = − 0.302). Two partial correlations (intervention partialized) showed a systematic impact of the type of exercise (LI-BFR vs. HI) (miR-99a-59: r = − 0.280/miR-199a-3p: r = − 0.293). Conclusion MiRNA expression patterns differ according to type of activity. We concluded that not only the intensity of the exercise (LC) is decisive for the release of circulating miRNAs—as essential is the type of training and the oxygen supply.

Myocardial infarction (MI) not only defines acute MI with obstructed coronary arteries (T1MI) but also myocardial necrosis caused by myocardial oxygen supply/demand mismatch as type 2 MI (T2MI); only T1MI patients benefit from an early invasive management. Myeloid-related protein(MRP)-8/14 is a biomarker described in various inflammatory diseases and in MI patients. Here we evaluate the potential of MRP-8/14 and high-sensitivity troponin I (hs-cTnI) to differentiate T2MI from T1MI. Patients with final diagnosis NSTEMI (n = 254; 33.1% female) enrolled in a prospective biomarker registry between 08/2011 and 10/2016 were analysed. Median baseline MRP-8/14 levels were higher in T2MI (n = 55; 3.37(1.88–6.48)μg/mL) than in T1MI (n = 199; 2.4 [1.4–3.79]μg/mL) (p = .013) patients, in contrast to hs-cTnI (T2MI:52[11.65–321.4]ng/L vs. T1MI:436.5 [61.25–1973.8]ng/L; p To detect the strength of this association odds ratios(OR) were calculated with MRP-8/14 yielding 2.13(1.16–3.92; p = .015) to predict T2MI and 0.47(0.26–0.87; p = .015) for T1MI. As expected, hs-cTnI yielded an OR of to predict T2MI 0.34(0.17–0.65; p = .001) and 2.98(1.53–5.81; p = .001) for T1MI. Both markers show comparable and independent results if adjust to hs-cTnI/MRP-8/14, TIMI risk score and CRP. T2MI is associated with higher MRP-8/14 and lower hs-cTnI concentrations than T1MI. Our data suggest that MRP-8/14 as a marker of inflammation might provide usable discriminatory information complementing hs-cTnI in a diagnostic procedure evaluating the type of MI directly upon hospital admission.

Introduction Although a number of risk factors for chronic thromboembolic pulmonary hypertension (CTEPH) have been reported, the exact prevalence is controversial and varies between published cohorts. The aim of the present study was to investigate the prevalence of risk factors in operable CTEPH patients with special emphasis on thyroid disease and function. Material and methods Overall, 228 CTEPH patients (47.7% female; median age 63 [IQR 52–72] years) scheduled for pulmonary endarterectomy between 01/2014 and 12/2015 were studied. Prevalence of risk factors was assessed, and patients were classified according to their thyroid function based on laboratory measurements. Results As many as 86.0% of patients reported a history of pulmonary embolism (PE; of those, 24.5% were diagnosed with “acute” PE less than six months before the diagnosis of CTEPH), 80.7% of patients had a blood group non-0 and 24.1% of patients had known thyroid disease (of those, 78.2% hypothyroidism). Laboratory measurements revealed thyroid dysfunction in 10.5% of patients (of those, 54.8% had no known thyroid disease). Patients with hypothyroid function had higher WHO functional classes, NT-proBNP levels and a lower cardiac index compared to patients with euthyroid function. Conclusions The prevalence of a history of PE and blood group non-0 was higher than previously reported. However, a relevant proportion of patients might have suffered from pre-existing CTEPH rather than acute PE. Thyroid disease and dysfunction were frequent and hypothyroid function associated with more severe disease.

The aim of this study was to evaluate the cartilage intermediate layer protein 1 (CILP1) as a biomarker of right ventricular dysfunction in patients with ischemic cardiomyopathy (ICM). CILP1 plasma concentrations were measured in 98 patients with ICM and 30 controls without any cardiac abnormalities. All participants underwent cardiac magnetic resonance imaging. Median CILP1 concentrations were higher in ICM than in controls. In the tertile analysis, low right ventricular ejection fraction (RVEF) and high right ventricular end-systolic volume index and N-terminal pro-brain natriuretic peptide (NT-proBNP) were associated with higher CILP1 levels in ICM. However, there were no associations between CILP1 concentrations and left ventricular (LV) parameters in this group. In receiver-operating characteristic (ROC) analysis CILP1 was a good predictor of RVEF 40% with an optimal cut-off value of 3545 pg/ml in ICM, whereas it was not predictive of LV ejection fraction (LVEF) 40% (area under the curve [AUC] = 0.57) There was no significant difference between the ROC curves of CILP1 (AUC = 0.72) and NT-proBNP (AUC = 0.77) for RVEF 40% (

Background Severe aortic stenosis (AS) is associated with left ventricular (LV) pressure overload that leads to myocardial remodelling and inflammatory processes. Interleukin 6 (IL6) is secreted by leukocytes as an early response to infection and tissue damage as well as high senisitve C-reactive Protein (hsCRP), which is subsequent in the same pathway. Several studies have suggested an association of elevated serum levels with a higher risk of cardiovascular events. GDF-15 and MR-proADM are also associated with inflammatory processes in cardiovascular diseases and are predictors for adverse events and mortality in patients with AS. The aim of the present study was to evaluate their potential prognostic value regarding the patients all-cause mortality. Methods A total of 92 consecutive patients (mean age: 80,8 [±5,3] years) undergoing TAVI were included in this study. TAVI was performed according to standard clinical practice. Venous blood samples for biomarker analysis were collected prior to and 6 months after TAVI, these were processed immediately and frozen at −80°C until the assay was performed. Safety events, physiological- and echocardiographical parameters, were assessed at the baseline and the 6-month follow-up. Furthermore, we compiled the all-cause mortality of our patients after two years. Results TAVI was performed successfully in all patients. During the two-year follow-up period 24 patients met the endpoint of all-cause mortality. At baseline, serum levels of the inflammatory biomarkers were significantly higher in patients who died within the follow-up period, when compared to survivors (IL6:14,450pg/ml [IQR:7,550; 42,150] vs. 4,200pg/ml [IQR:2,515; 13,875],p=0,0004; hsCRP:5,360 mg/l [IQR:2,248; 26,790] vs. 2,900mg/l [IQR:1,208; 8,210],p=0,022); MR-proADM:1,347nmol/l [IQR:1,038–1,678] vs. 0,922nmol/l [IQR:0,706; 1,202],p=0,0003 and GDF-15:2770,0pg/ml [IQR:2401,0; 3701,0] vs. 1675,2pg/ml [IQR:1141,6; 2524,4],p=0,001). The area under the curve was 0,767 for IL-6, 0,665 for hsCRP, 0,735 for MR-proADM and 0,735 for GDF-15. In addition, there was a significant decrease of IL-6 (baseline: 4,200pg/ml [IQR:2,525; 13,875] vs. 6FU:2,600pg/ml [IQR:1,500; 7,000],p Conclusions In the present study there was a significant decrease of inflammatory biomarkers after TAVI in high risk patients with severe aortic stenosis and good clinical outcome. In this regard, IL-6, hsCRP, MR-proADM and GDF-15 were predictors of all-cause mortality in patients, who underwent TAVI. Funding Acknowledgement Type of funding sources: None.

Background Walking is the preferred therapy for peripheral arterial disease in early stage. An effect of walking exercise is the increase of blood flow and fluid shear stress, leading, triggered by arteriogenesis, to the formation of collateral blood vessels. Circulating micro-RNA may act as an important information transmitter in this process. We investigated the acute effects of a single bout of 1) aerobic walking with moderate intensity; and 2) anaerobic walking with vigorous intensity on miRNA parameters related to vascular collateral formation. Methods Ten (10) patients with peripheral arterial disease with claudication (age 72 ± 7 years) participated in this two-armed, randomized-balanced cross-over study. The intervention arms were single bouts of supervised walking training at (1) vigorous intensity on a treadmill up to volitional exhaustion and (2) moderate intensity with individual selected speed for a duration of 20 min. One week of washout was maintained between the arms. During each intervention, heart rate was continuously monitored. Acute effects on circulating miRNAs and lactate concentration were determined using pre- and post-intervention measurement comparisons. Results Vigorous-intensity walking resulted in a higher heart rate (125 ± 21 bpm) than the moderate-intensity intervention (88 ± 9 bpm) (p p = 0.005; 3.3 ± 1.2 mmol/l), but not after moderate exercising (p > 0.05; 1.7 ± 0.6 mmol/l). The circulating levels of miR-142-5p and miR-424-5p were up-regulated after moderate-intensity (p p > 0.05). Conclusion Moderate-intensity walking seems to be more feasible than vigorous exercises to induce changes of blood flow and endurance training-related miRNAs in patients with peripheral arterial disease. Our data thus indicates that effect mechanisms might follow an optimal rather than a maximal dose response relation. Steady state walking without the necessity to reach exhaustion seems to be better suited as stimulus.

Aim: This study assessed the utility of osteopontin (OPN) and galectin-3 (Gal-3) as biomarkers of maladaptive right ventricular remodeling in pulmonary hypertension (PH). Materials & methods: We examined plasma levels of OPN and Gal-3 in patients with PH (n = 62), dilated cardiomyopathy (n = 34), left ventricular hypertrophy (LVH; n = 47), and controls without right ventricle (RV) or LV abnormalities (n = 38). Results: OPN and Gal-3 levels were higher in PH, dilated cardiomyopathy and LVH than in the controls. OPN concentrations in PH patients with maladaptive RV were significantly higher than in those with adaptive RV. Gal-3 did not differentiate between adaptive and maladaptive RV remodeling in PH. OPN and Gal-3 levels did not correlate with parameters of LV remodeling. Conclusion: OPN is a potential biomarker of RV maladaptation.

Aims Disturbance of mitochondrial function significantly contributes to the myocardial injury that occurs during reperfusion. Increasing evidence suggests a role of intra-mitochondrial cyclic AMP (cAMP) signaling in promoting respiration and ATP synthesis. Mitochondrial levels of cAMP are controlled by type 10 soluble adenylyl cyclase (sAC) and phosphodiesterase 2 (PDE2), however their role in the reperfusion-induced injury remains unknown. Here we aimed to examine whether sAC may support cardiomyocyte survival during reperfusion. Methods and results Adult rat cardiomyocytes or rat cardiac H9C2 cells were subjected to metabolic inhibition and recovery as a model of simulated ischemia and reperfusion. Cytosolic Ca2+, pH, mitochondrial cAMP (live-cell imaging), and cell viability were analyzed during a 15-min period of reperfusion. Suppression of sAC activity in cardiomyocytes and H9C2 cells, either by sAC knockdown, by pharmacological inhibition or by withdrawal of bicarbonate, a natural sAC activator, compromised cell viability and recovery of cytosolic Ca2+ homeostasis during reperfusion. Contrariwise, overexpression of mitochondria-targeted sAC in H9C2 cells suppressed reperfusion-induced cell death. Analyzing cAMP concentration in mitochondrial matrix we found that inhibition of PDE2, a predominant mitochondria-localized PDE isoform in mammals, during reperfusion significantly increased cAMP level in mitochondrial matrix, but not in cytosol. Accordingly, PDE2 inhibition attenuated reperfusion-induced cardiomyocyte death and improved recovery of the cytosolic Ca2+ homeostasis. Conclusion sAC plays an essential role in supporting cardiomyocytes viability during reperfusion. Elevation of mitochondrial cAMP pool either by sAC overexpression or by PDE2 inhibition beneficially affects cardiomyocyte survival during reperfusion.

Coronary artery disease (CAD) is a long-lasting inflammatory disease characterized by monocyte migration into the vessel wall leading to clinical events like myocardial infarction (MI). However, the role of monocyte subsets, especially their miRNA-driven differentiation in this scenario is still in its infancy. Here, we characterized monocyte subsets in controls and disease phenotypes of CAD and MI patients using flow cytometry and miRNA and mRNA expression profiling using RNA sequencing. We observed major differences in the miRNA profiles between the classical (CD14

Background Chronic thromboembolic pulmonary hypertension (CTEPH) is a sub group of pulmonary hypertension (PH). CTEPH is characterized by the existence of thromboemboli and vascular remodeling in pulmonary vessels. The effect of increase in pulmonary artery pressures causes right ventricle (RV) hypertrophy and dilatation and finally leads to right heart failure and death. Surgical intervention in operable patients makes the CTEPH as an only curable and unique form of ph. Pulmonary endarterectomy (PEA) is the surgical procedure to remove the thromboembolic clots from the pulmonary vasculature, which restores RV function back to normal with significant improvements in cardiovascular magnetic resonance. Purpose The aim of this study is to use transcriptomic profiling to identify signaling pathways, master regulators, and potentially new biomarkers that specifically indicate the effect of PEA on the RV of patients with chronic thromboembolic pulmonary hypertension. Results RNA -sequencing (RNA-seq) was performed on RV biopsies obtained from CTEPH patients at PEA baseline (before PEA surgery) and the results were compared with those from RV biopsies obtained during follow-up evaluation. Bioinformatic analysis of RNA-seq data identified 2799 genes (n=14, −0.585 ≤ Log2 fold change ≥0.585, FDR ≤0.05) differentially regulated between the PEA baseline and follow-up sample groups. The great number of genes (2799) differentially expressed after PEA surgery in CTEPH patients confirms a major shift in the transcriptional landscape of RV in these patients. To further identify potential biomarker candidates from the large pool of 2799 differentially expressed genes (DEGs), extensive bioinformatic analysis of different data sets shortlisted 250 DEGs that were functionally associated with cardiovascular development or disease. The findings of this study reveal prominent transcriptional changes that occur in response to PEA. Gene ontology enrichment and pathway analysis confirmed altered regulation of hypoxia-inducible factor 1 (HIF-1) signaling, advanced glycation end products and their receptors (AGE-RAGE), mitogen-activated protein kinase (MAPK) signaling, hippo signaling, the Janus kinase/ signal transducers and activators of transcription (Jak-STAT) signaling pathway, and proteoglycans after PEA compared with before PEA. Conclusion Comparison of the results of RNA-seq analysis of RV biopsies of CTEPH patients, pre and post PEA, revealed a major shift in the transcriptional landscape of these patients after reducing the pressure overload of the RV by PEA. Funding Acknowledgement Type of funding source: Foundation. Main funding source(s): German Research Foundation (DFG)

Background Cardiac troponins have become the gold standard for appropriate and rapid diagnosis of an acute myocardial infarction (AMI). However, elevated cardiac troponins levels can be observed in the absence of AMI in elderly patients and therefore a loss of specificity is seen in the diagnosis of AMI in elderly patients. Purpose This study aims to define old age in suspected AMI and to evaluate the effect of older age on the diagnostic specificity of a high sensitivity troponin I (hs-cTnI) and high sensitivity troponin T (hs-cTnT) assay in elderly women and men. Methods This study used data from patients with suspected AMI who were enrolled between 08–2011 and 10–2016 to a multicentre biomarker registry and an University Hospital and with available hs-cTnI and hs-cTnT upon admission (n=564 patients). First, we investigated the effect of age on the specificity of both hs-cTnI and hs-cTnT to detect AMI continuously. We then classified over seventy-year-olds as elderly (n=281; 49,8%) and compared the specificity of hs-cTnI and hs-cTnT between elderly and younger patients. Results In the group of older patients, 62.6% (n=176) were diagnosed with AMI compared with 50.9% (n=144) in younger age patients (p Conclusions The specificity of both, hs-cTnI and hs-cTnT is significantly reduced in elderly patients due to higher hs-cTnT and hs-cTnI levels in elderly patients at admission irrespective of the final diagnosis. A relevant loss in diagnostic specificity is observed at 70 years of age starting already at 60 years. Use of assay-specific adjusted cut-offs for hs-cTnI and hs-cTnT for elderly patients regains diagnostic specificity that leads to a more accurate decision-making concerning “rule-in” for older patients with suspected AMI. Funding Acknowledgement Type of funding source: None

Arteriogenesis is a process by which a pre-existing arterioarterial anastomosis develops into a functional collateral network following an arterial occlusion. Alternatively activated macrophages polarized by IL10 have been described to promote collateral growth. This study investigates the effect of different levels of IL10 on hind-limb reperfusion and the distribution of perivascular macrophage activation types in mice after femoral artery ligation (FAL). IL10 and anti-IL10 were administered before FAL and the arteriogenic response was measured by Laser-Doppler-Imaging perioperatively, after 3, 7, and 14 d. Reperfusion recovery was accelerated when treated with IL10 and impaired with anti-IL10. Furthermore, symptoms of ischemia on ligated hind-limbs had the highest incidence after application of anti-IL10. Perivascular macrophages were immunohistologically phenotyped using CD163 and CD68 in adductor muscle segments. The proportion of alternatively activated macrophages (CD163+/CD68+) in relation to classically activated macrophages (CD163&minus, /CD68+) observed was the highest when treated with IL10 and suppressed with anti-IL10. This study underlines the proarteriogenic response with increased levels of IL10 and demonstrates an in-vivo alteration of macrophage activation types in the perivascular bed of growing collaterals.

Objective: Arteriogenesis, describing the process of collateral artery growth, is activated by fluid shear stress (FSS). Since this vascular mechanotransduction may involve microRNAs (miRNAs), we investigated the FSS-induced expression of vascular cell miRNAs and their functional impact on collateral artery growth during arteriogenesis. Approach and Results: To this end, rats underwent femoral artery ligation and arteriovenous anastomosis to increase collateral blood flow to maximize FSS and trigger collateral vessel remodeling. Five days after surgery, a miRNA expression profile was obtained from collateral tissue, and upregulation of 4 miRNAs (miR-24-3p, miR-143-3p, miR-146a-5p, and miR-195-5p) was verified by quantitative polymerase chain reaction. Knockdown of miRNAs at the same time of the surgery in an in vivo mouse ligation and recovery model demonstrated that inhibition of miR-143-3p only severely impaired blood flow recovery due to decreased arteriogenesis. In situ hybridization revealed distinct localization of miR-143-3p in the vessel wall of growing collateral arteries predominantly in smooth muscle cells. To investigate the mechanotransduction of FSS leading to the increased miR-143-3p expression, cultured endothelial cells were exposed to FSS. This provoked the expression and release of TGF-β (transforming growth factor-β), which increased the expression of miR-143-3p in smooth muscle cells in the presence of SRF (serum response factor) and myocardin. COL5A2 (collagen type V-α2)—a target gene of miR-143-3p predicted by in silico analysis—was found to be downregulated in growing collaterals. Conclusions: These results indicate that the increased miR-143-3p expression in response to FSS might contribute to the reorganization of the extracellular matrix, which is important for vascular remodeling processes, by inhibiting collagen V-α2 biosynthesis.

Aim: This study assessed the utility of SPARC-like protein 1 (SPARCL1) as a biomarker of maladaptive right ventricular (RV) function in patients with pulmonary hypertension (PH). Methods: In this prospective study, we examined SPARCL1 levels in 105 patients with adaptive (n = 34) and maladaptive RV (n = 32) pressure overload caused by PH, dilated cardiomyopathy (DCM, n = 18) with LVEF < 35% and preserved RV function and controls without LV or RV abnormalities (n = 21). Results: The median SPARCL1 concentration in patients with maladaptive RV function was higher than in those with adaptive RV function (p p p p p Conclusion: SPARCL1 shows potential as novel biomarker of RV pathological remodelling and is associated with RV maladaptation and ventriculoarterial uncoupling in PH.

BACKGROUND: Compromised renal function is a major risk factor that is strongly associated with poor outcome in patients with mitral regurgitation (MR) and heart failure. Cystatin C, a cysteine protease inhibitor, has been used as a specific and sensitive biomarker of renal function. Neutrophil gelatinase‐associated lipocalin (NGAL) is another sensitive biomarker that specifically indicates functional and structural kidney damage. The aim of the present study was to determine the predictive value of serum cystatin C and urinary NGAL as indicators of mortality in patients undergoing percutaneous mitral valve repair (PMVR). METHODS: A total of 120 consecutive patients (age: 77.3 years [±11.2]) undergoing PMVR using the MitraClip system were included in this study. Venous blood and urinary samples were collected for biomarker analysis prior to PMVR. Physiological parameters, medication use, safety events, and all‐cause mortality were assessed 12 months after the procedure. RESULTS: Twelve months after PMVR, there was a significant reduction in the severity of MR (P < 0.001), and an improvement in the New York Heart Association class (P < 0.01) was documented. Baseline levels of serum cystatin C (nonsurvivors: 2.4 mg/L [interquartile, IQR: 1.7;3.1] vs survivors: 1.7 mg/L [IQR: 1,3;2.1], P < 0.001) and urinary NGAL (nonsurvivors: 242.0 ng/mL [IQR: 154.5;281.5] vs survivors: 132.0 ng/mL [IQR:107.0;177.3], P < 0.001) were significantly higher in patients who died during the 12‐month follow‐up period. CONCLUSION: Cystatin C and urinary NGAL were found to be predictors of long‐term mortality in high‐risk patients undergoing PMVR. Thus, cystatin C and NGAL assessment may be helpful in risk stratification in patients undergoing PMVR.

Objective Blood monocyte subsets are emerging as biomarkers of cardiovascular inflammation. However, our understanding of human monocyte heterogeneity and their immunophenotypic features under healthy and inflammatory conditions is still evolving. Rationale In this study, we sought to investigate the immunophenome of circulating human monocyte subsets. Methods Multiplexed, high-throughput flow cytometry screening arrays and computational data analysis were used to analyze the expression and hierarchical relationships of 242 specific surface markers on circulating classical (CD14++CD16−), intermediate (CD14++CD16+), and nonclassical (CD14+CD16++) monocytes in healthy adults. Results Using generalized linear models and hierarchical cluster analysis, we selected and clustered epitopes that most reliably differentiate between monocyte subsets. We validated existing transcriptional profiling data and revealed potential new surface markers that uniquely define the classical (e.g., BLTR1, CD35, CD38, CD49e, CD89, CD96), intermediate (e.g., CD39, CD275, CD305, CDw328), and nonclassical (e.g., CD29, CD132) subsets. In addition, our analysis revealed phenotypic cell clusters, identified by dendritic markers CMRF-44 and CMRF-56, independent of the traditional monocyte classification. Conclusion These results reveal an advancement of the clinically applicable multiplexed screening arrays that may facilitate monocyte subset characterization and cytometry-based biomarker selection in various inflammatory disorders.

Background An association between Tako-Tsubo cardiomyopathy (TTC) and underlying malignancies has been observed, suggesting that TTC might be the consequence of paraneoplastic phenomena. This study investigates the presence of autoantibodies against cardiomyocytes as well as adrenergic (β1, β2) and muscarinic (M2) receptors in patients with TTC. Methods and results Serum from 20 TTC patients and 20 controls with ischemic heart disease was obtained. Indirect immunofluorescence testing for intracellular autoantibodies against cardiomyocytes showed a homogenous distribution, as in both groups 9 of 20 sera displayed a characteristic binding pattern of antibodies including vascular walls and intracellular structures. Flow cytometry analysis revealed no difference between TTC and controls in the binding of autoantibodies to the surface antigens of cardiomyocyte HL-1 cells (p = 0.569, t-test). Flow cytometry analysis of nontransfected wild type cells (p = 0.633, t-test), M2 receptor-transfected cells (p = 0.687, t-test), β1 receptor-transfected cells (p = 0.444, t-test) and β2 receptor-transfected cells (p = 0.632, t-test) showed similar results for control and TTC sera. Likewise, the binding pattern of TTC patients with a history of neoplasia compared to those without or to controls did not differ significantly (p > 0.05, u-test). Conclusion Findings suggest that the presumed paraneoplastic etiology of TTC cannot be attributed to the formation of these antibodies.

Cardiomyopathies are a leading cause of heart failure and are often caused by mutations in sarcomeric genes, resulting in contractile dysfunction and cellular damage. This may stimulate the production of a robust proinflammatory response. To determine whether myocardial inflammation is associated with cardiac dysfunction in dilated cardiomyopathy (DCM) caused by MYBPC3 mutation, we used the well-characterized cMyBP-C(t/t) mouse model of DCM at 3 months of age. Compared to wild type (WT) mice, DCM mice exhibited significantly decreased fractional shortening (36.4±2% vs. 15.5±1.0%, p < 0.0001) and significantly increased spleen weight (5.3±0.3% vs. 7.2±0.4 mm/mg, p = 0.002). Intriguingly, flow cytometry analysis revealed a significant increase in total (CD45+CD11b+Ly6C−MHCII+F480+) macrophages (6.5±1.4% vs. 14.8±1.4%, p = 0.002) and classically activated (CD45+CD11b+Ly6C−MHCII+F480+CD206−) proinflammatory (M1) macrophages (3.4±0.8% vs. 10.3±1.2%, p = 0.0009) in DCM hearts as compared with WT hearts. These results were further confirmed by immunofluorescence analysis of heart tissue sections. Splenic red pulp (CD11b+Ly6C+MHCIIlowF480hi) macrophages were significantly elevated (2.4±0.1% vs. 1.3±0.1%, p = 0.0001) in DCM compared to WT animals. Serum cytokine analysis in DCM animals exhibited a significant increase (0.65±0.2 vs. 2.175±0.5 pg/mL, p = 0.02) in interleukin (IL)-6 compared to WT animals. Furthermore, RNAseq analysis revealed the upregulation of inflammatory pathways in the DCM hearts. Together, these data indicate a robust proinflammatory response in DCM hearts, likely in response to cellular damage triggered by MYBPC3 mutation and resultant contractile dysfunction.

Extracellular vesicles are released by numerous cell types of the human body under physiological but also under pathophysiological conditions. They are important for cell&ndash, cell communication and carry specific signatures of peptides and RNAs. In this study, we aimed to determine whether extracellular vesicles isolated from patients with pulmonary hypertension show a disease specific signature of small non-coding RNAs and thus have the potential to serve as diagnostic and prognostic biomarkers. Extracellular vesicles were isolated from the serum of 23 patients with chronic thromboembolic pulmonary hypertension (CTEPH) and 23 controls using two individual methods: a column-based method or by precipitation. Extracellular vesicle- associated RNAs were analyzed by next-generation sequencing applying molecular barcoding, and differentially expressed small non-coding RNAs were validated by quantitative real-time polymerase chain reaction (qRT-PCR). We identified 18 microRNAs and 21 P-element induced wimpy testis (PIWI)-interacting RNAs (piRNAs) or piRNA clusters that were differentially expressed in CTEPH patients compared with controls. Bioinformatic analysis predicted a contribution of these piRNAs to the progression of cardiac and vascular remodeling. Expression levels of DQ593039 correlated with clinically meaningful parameters such as mean pulmonary arterial pressure, pulmonary vascular resistance, right ventricular systolic pressure, and levels of N-terminal pro-brain natriuretic peptide. Thus, we identified the extracellular vesicle- derived piRNA, DQ593039, as a potential biomarker for pulmonary hypertension and right heart disease.

Aims and background Hypertension is a disease accompanied by moderate inflammation. Physical activity and pharmacological intervention protect the organisms against hypertension-dependent end-organ damage. They should therefore interfere with the mobilization of lymphocytes. Different subsets of lymphocytes should specifically interfere with vascular and cardiac remodeling. In this study we correlated the amount of circulating lymphocytes with the expression of vascular and cardiac specific genes in the aorta and left ventricle to identify the tissue-specific effects of these cells. Methods 50 female spontaneously hypertensive rats (SHR) and 6 female normotensive Wistar rats (Wis) were analyzed. They were grouped in 7 subgroups exposed to different treatment regimes (modification of activity, type of activity, aldosterone blockade, and a combination thereof. Indicator genes of mal-adaptive vascular and cardiac remodeling were quantified by qRT-PCR. Results SHRs had an increased number of circulating NK cells and monocytes, but lower numbers of T cells. T cells inversely correlated with the vascular expression of p22phox, a member of the NADPH oxidase complex. Thus, lower number of T-cells favors the vascular expression of p22phox and thereby oxidative stress. In the left ventricle, T cells are inversely correlated with the expression of arginase-1 and UCP-2 but positively correlated with Glut-4 expression. The lower number of T cells in SHR therefore favors the expression of proteins often associated with oxidative stress and glucose metabolism. Monocytes are positively correlated with key markers of left ventricular hypertrophy (such as ANP, collagen-1, and β-MHC). Thus, the increased number of monocytes in SHRs supports cardiac hypertrophy. NK cells are inversely correlated with the expression of anti-inflammatory cytokines in the aorta (IL-10, IL-15). Thus, high number of circulating NK cells seems to favor a pro-inflammatory phenotype in the vasculature. Conclusion In a standard model of essential hypertension the profile of circulating lymphocytes differs from that in normotensive rats. The different subsets of lymphocytes specifically interfere with specific processes in the ventricle and vessel. Collectively, the difference between hypertensive and normotensive rats (less T cells, more NK cells and monocytes) supports a mal-adaptive role under hypertensive conditions.

Background Acute myocardial infarction (MI) is associated with high morbidity and mortality. A robust differentiation between type 1 and type 2 MI (T1/T2MI) has prognostic and therapeutic implications. We investigated whether serial high-sensitivity cardiac troponin I measurements could reliably discriminate T1MI from T2MI in patients presenting with a non-ST elevation myocardial infarction (NSTEMI). Methods We used data from a prospective acute coronary syndrome biomarker registry of patients with suspected MI that presented at or were transferred to one of two study centres. Here, we analysed an unselected group of 265 NSTEMI patients (67.2% males). Blood was drawn on admission and after 3 hours. High-sensitivity troponin I (hs-cTnI) was measured in frozen samples by a technician blinded to patient characteristics. T1MI or T2MI was defined as the gold-standard study diagnosis by two independent cardiologists based on all available data according to the Third Universal Definition of MI. Results A diagnosis of T2MI was made in 55 patients (20.8%) in the NSTEMI cohort. T2MI patients did not differ from T1MI patients regarding age, gender, traditional risk factors, or percentage of those with a history of coronary artery disease. Median baseline hs-cTnI levels were higher in T1MI (436.25; IQR 63.7–1918.8 ng/L) than in T2MI patients (48.4; IQR 11.7–305.9 ng/L; p Conclusion Our data show that hs-cTnI concentrations differ between patients presenting with T1 and T2MI. The concentration of hs-cTnI and its change over time has the potential to rule out T2MI and therefore to identify patients who might benefit from an early invasive management. The differentiation between T1MI and T2MI by using hs-cTnI is nevertheless challenging, and further research on specific algorithms is needed. Acknowledgement/Funding 3German Center for Cardiovascular Research (DZHK), Partnersite Rhein Main, Bad Nauheim, Germany

Aims and background Spontaneously hypertensive rats (SHR) are a suitable model of essential hypertension and allow analyze the progression of hypertension and hypertension-dependent end-organ damage. In this model, acute improvement of physical activity (free running wheel activity) exerts known beneficial effects (such as lowering oxidative stress). However, these initial beneficial effects are lost during the continuation of high physical activity and translate into mal-adaptive processes, suggesting that not any high physical activity exerts beneficial effects. It has been hypothesized that the skeletal muscle release myokines, that contribute to the beneficial effects of exercise. However, myokines, such as IL-6, are induced by an acute increase in work load not by continuous work load. Therefore, we analyzed whether modification of high physical activity, i.e. intermittent free running wheel activity, modify the long-term impact on long lasting hypertension. Methods 38 female SHR aged 6 weeks (pre-hypertensive state) were randomly allocated to one of the following groups: Sedentary (S; standard holding condition) imitating the condition of sedentary life style, high activity (HA; life-long free running wheel) imitating the condition of active life style, temporary activity (TA; 6 months free running wheel and 3 months sedentary) imitating the loss of active life style during ageing, and finally intermittent activity (IA; 10 months repetitive access to running wheels every 4 weeks) imitating altered workloads. All rats were sacrificed at the age of 10 months. Results IA was the only treatment regime that effectively lowered blood pressure (P syst: 186±11 vs. 165±6 mmHg), improved ejection fraction (EF: 56±5 vs. 63±2%), and displayed clear molecular profile of adaptive myocardial hypertrophy rather than mal-adaptive hypertrophy. Moreover, only IA reduced the number of circulating monocytes (377±102 vs. 220±16 /μl), a cell population that immigrated in the left ventricle. The number of monocytes was directly correlated with the expression of MMP12, BNP, ANP, biglycan, collagen-1, actinin, β-MHC, and somatstatin but inversely related to β-adrenoceptor, Glut-4, and UCP3 expression. Finally, IA increased the skeletal expression of IL-6 and decreased the renal expression of AT1 receptors. Conclusion The data confirm the previous findings that not all type of physical activity beneficially affects hypertensive-dependent disease. In contrast, the data support the hypothesis that alterations in work load are required triggering the release of myokines from the skeletal muscle and identify the amount of circulating monocytes as a main trigger of mal-adaptive hypertrophy in these rats. The data are important with respect to optimize life style suggestions for patients with essential hypertension. Acknowledgement/Funding DFG (ERAGON and SFB 1213)

Background There are several tools for primary prevention (e.g. Framingham, ESC) that can be used to predict mortality risk in healthy individuals. However, only a few scores have been validated to predict outcome in patients with cardiovascular disease. One of these instruments is the REACH (REduction of Atherothrombosis for Continued Health) score. The ESC guideline for stable coronary artery disease (CAD) places a clear emphasis on carrying out risk stratification before using invasive treatment. Recent studies have revealed a prognostic value of serum hs-cTnI in patients with stable CAD. Purpose The aim of this study was to evaluate the prognostic information provided by hs-cTnI in stable high-risk CAD patients. Methods Between 2011 and 2014, consecutive stable patients with suspected CAD undergoing coronary angiography were included in the study. Data from a 4-year follow-up was obtained; the study endpoint was defined as all-cause mortality. Serum hs-cTnI was measured before angiography using a high-sensitivity assay. Results A total of 3,742 patients were included, of whom 2,274 (60.1%) had confirmed CAD. Patients with an estimated annual mortality rate above 3% using the REACH score were defined as having high risk (n=996 in the low-risk group, n=1,278 in the high-risk cohort). Patients with higher risk were more often male (81.5% vs. 69.2%, p Conclusion Hs-cTnI was found to be an independent risk factor in low- as well as high-risk patients. Hs-cTnI levels correlate with the REACH risk score. Moreover, it was possible to separate patients at very high and very low risk by combining REACH score and hs-cTnI.

Introduction and aim Vitamin D deficiency is associated with an adverse prognosis in patients with coronary artery disease (CAD). Decreased levels of vitamin D are associated with low sunshine exposure, resulting in seasonal variations of vitamin D. The aims of this study were to investigate the influence of different specific weather conditions on vitamin D levels and to explore a possible improvement of risk stratification by vitamin D levels in stable patients with CAD using meteorological data. Methods The study population consists of two independent cohorts of stable patients undergoing coronary angiography with suspected or known CAD: as derivation cohort, the ongoing biomarker registry BioPROSPECTIVE with n=1,766 enrolled patients between 2010 and 2013 (median age 70.1 yrs; 30.8% females); and as validation cohort, the Ludwigshafen Risk and Cardiovascular Health (LURIC) Study with n=3,299 patients (median age 63.5 yrs; 30.3% females). In the derivation cohort 235 (13.3%) patients were known to be deceased by 08/2018. In the validation cohort 760 (23.0%) patients died within a median follow-up time of 7.75 years. 25-OH vitamin D levels were measured by commercial assays. Vitamin D deficiency was defined as 25-OH vitamin D levels ≤20 ng/mL. Daily averaged data on six weather conditions of the 180 days prior to enrolment were collected for each patient from the weather station located closest to the respective study centre. Using air pressure, precipitation height, sunshine duration, temperature, relative humidity, and vapour pressure a weather model was constructed that significantly correlated with vitamin D levels (r=0.37; p Results In the derivation cohort, median vitamin D levels were lower in non-survivors (13.3 [9.65–19.65] ng/mL) than in survivors (15.70 [10.7–22.65] ng/mL; p Conclusions Different weather conditions show a significant impact on vitamin D levels in stable patients. Adding data on weather conditions improve the risk stratification by vitamin D for predicting mortality in stable CAD patients. Acknowledgement/Funding The study is financially supported by the Kerckhoff Heart Research Institute (KHFI) and the German Center for Cardiovascular Research (DZHK).

Purpose: To evaluate vascular function and its relationship to cardiorespiratory fitness in professional handball athletes. Method: We examined 30 male professional handball athletes (age 2...

Rationale: Activated cardiac fibroblasts (CF) are crucial players in the cardiac damage response; excess fibrosis, however, may result in myocardial stiffening and heart failure development. Inhibition of activated CF has been suggested as a therapeutic strategy in cardiac disease, but whether this truly improves cardiac function is unclear. Objective: To study the effect of CF ablation on cardiac remodeling. Methods and Results: We characterized subgroups of murine CF by single-cell expression analysis and identified periostin as the marker showing the highest correlation to an activated CF phenotype. We generated bacterial artificial chromosome–transgenic mice allowing tamoxifen-inducible Cre expression in periostin-positive cells as well as their diphtheria toxin-mediated ablation. In the healthy heart, periostin expression was restricted to valvular fibroblasts; ablation of this population did not affect cardiac function. After chronic angiotensin II exposure, ablation of activated CF resulted in significantly reduced cardiac fibrosis and improved cardiac function. After myocardial infarction, ablation of periostin-expressing CF resulted in reduced fibrosis without compromising scar stability, and cardiac function was significantly improved. Single-cell transcriptional analysis revealed reduced CF activation but increased expression of prohypertrophic factors in cardiac macrophages and cardiomyocytes, resulting in localized cardiomyocyte hypertrophy. Conclusions: Modulation of the activated CF population is a promising approach to prevent adverse cardiac remodeling in response to angiotensin II and after myocardial infarction.

A total of 90 consecutive patients undergoing renal sympathetic denervation (RSD) were included in this study. A significant reduction in office systolic blood pressure (SBP) of 21.1 mm Hg (P

Previous studies of renal sympathetic denervation (RSD) excluded patients with impaired renal function to avoid potential RSD-related renal damage. Measurement of the highly sensitive biomarkers neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule-1 (KIM-1) has shown that RSD does not aggravate renal damage during the early post-procedural period. The aim of the present study was to examine the effect of RSD on blood pressure (BP) reduction and renal function after a long-term follow-up. A total of 62 consecutive patients undergoing RSD were included in this study. Serum NGAL and KIM-1 were collected prior to RSD and at 24 hr, 48 hr, and 3 months after RSD. BP measurements, antihypertensive medication use, and safety events were followed over a three-year period. Follow-up data were available over 36.9[±3.4] months in 47 of 62 (75.8%) of the initially included patients. At this time point a significant systolic BP reduction of 23 mm Hg (P > 0.001) was documented, and there were no significant changes in serum creatinine (P = 0.14), blood urea nitrogen (P = 0.33), or estimated glomerular filtration rate (eGFR) (P = 0.2) values. There were also no significant changes documented in patients with impaired renal function (eGFR

The aim of our study was to analyse the protein expression of cartilage intermediate layer protein (CILP)1 in a mouse model of right ventricular (RV) pressure overload and to evaluate CILP1 as a biomarker of cardiac remodelling and maladaptive RV function in patients with pulmonary hypertension (PH).Pulmonary artery banding was performed in 14 mice; another nine mice underwent sham surgery. CILP1 protein expression was analysed in all hearts using Western blotting and immunostaining. CILP1 serum concentrations were measured in 161 patients (97 with adaptive and maladaptive RV pressure overload caused by PH; 25 with left ventricular (LV) hypertrophy; 20 with dilative cardiomyopathy (DCM); 19 controls without LV or RV abnormalities)In mice, the amount of RV CILP1 was markedly higher after banding than after sham. Control patients had lower CILP1 serum levels than all other groups (p−1) was associated with lower tricuspid annular plane excursion/pulmonary artery systolic pressure ratios (pCILP1 is a novel biomarker of RV and LV pathological remodelling that is associated with RV maladaptation and ventriculoarterial uncoupling in patients with PH.

BACKGROUND: Percutaneous mitral valve repair (PMVR) is an interventional treatment option in patients with severe mitral regurgitation (MR) and at high risk for open‐heart surgery. Currently, limited information exists about predictors of procedural success after PMVR. Galectin‐3 (Gal‐3) and suppression of tumorigenicity 2 (ST2) induce fibrotic alterations in severe MR and heart failure. We sought to examine the predictive value of Gal‐3 and ST2 as specific indicators of therapeutic success in high‐risk patients undergoing PMVR. HYPOTHESIS: We hypothesize that extended cardiac fibrotic alterations might have impact on successful MR reduction after the MitraClip procedure. METHODS: A total of 210 consecutive patients undergoing PMVR using the MitraClip system were included in this study. Procedural success was defined as an immediate reduction of MR by ≥2 grades, assessed by echocardiography. Venous blood samples were collected prior to PMVR and at 6 months follow‐up for biomarker analysis. RESULTS: After PMVR there was a significant reduction in the severity of MR (MR grade: 3 ±0.3 vs 1.6 ±0.6, P