The role of Matrix Metalloproteinase-2 and Galectin-3 as predictive biomarkers for all-cause mortality in patients undergoing transfemoral transcatheter aortic valve implantation.

BACKGROUND: Currently available risk scores fail to accurately predict morbidity and mortality in patients with severe symptomatic aortic stenosis who undergo transcatheter aortic valve implantation (TAVI). In this context, biomarkers like matrix metalloproteinase-2 (MMP-2) and Galectin-3 (Gal-3) may provide additional prognostic information. METHODS: Patients with severe aortic stenosis undergoing consecutive, elective, transfemoral TAVI were included. Baseline demographic data, functional status, echocardiographic findings, clinical outcomes and biomarker levels were collected and analysed. RESULTS: The study cohort consisted of 89 patients (age 80.4 ± 5.1 years, EuroScore II 7.1 ± 5.8%). During a median follow-up period of 526 d, 28 patients (31.4%) died. Among those who died, median baseline MMP-2 (alive: 221.6 [170.4; 263] pg/mL vs. deceased: 272.1 [225; 308.8] pg/mL, p < 0.001) and Gal-3 levels (alive: 19.1 [13.5; 24.6] pg/mL vs. deceased: 25 [17.6; 29.5] pg/mL, p = 0.006) were higher than in survivors. In ROC analysis, MMP-2 reached an acceptable level of discrimination to predict mortality (AUC 0.733, 95% CI [0.62; 0.83], p < 0.001), but the predictive value of Gal-3 was poor (AUC 0.677, 95% CI [0.56; 0.79], p = 0.002). Kaplan-Meier and Cox regression analyses showed that patients with MMP-2 and Gal-3 concentrations above the median at baseline had significantly impaired long-term survival (p = 0.004 and p = 0.02, respectively). CONCLUSIONS: In patients with severe aortic stenosis undergoing transfemoral TAVI, MMP-2 and to a lesser extent Gal-3, seem to have additive value in optimizing risk prediction and streamlining decision-making.

Changes in miRNA expression in patients with peripheral arterial vascular disease during moderate- and vigorous-intensity physical activity.

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 < 0.05). Lactate concentration was increased after vigorous-intensity walking (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 < 0.05), but not after vigorous-intensity training (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.

CILP1 as a biomarker for right ventricular maladaptation in pulmonary hypertension.

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<0.001). CILP1 concentrations were higher in PH patients with maladaptive RV function than those with adaptive RV function (p<0.001), LV pressure overload (p<0.001) and DCM (p=0.003). CILP1 showed good predictive power for maladaptive RV in receiver operating characteristic analysis (area under the curve (AUC) 0.79). There was no significant difference between the AUCs of CILP1 and N-terminal pro-brain natriuretic peptide (NT-proBNP) (AUC 0.82). High CILP1 (cut-off value for maladaptive RV of ≥4373 pg·mL-1) was associated with lower tricuspid annular plane excursion/pulmonary artery systolic pressure ratios (p<0.001) and higher NT-proBNP levels (p<0.001).CILP1 is a novel biomarker of RV and LV pathological remodelling that is associated with RV maladaptation and ventriculoarterial uncoupling in patients with PH.

Shear Stress-Induced miR-143-3p in Collateral Arteries Contributes to Outward Vessel Growth by Targeting Collagen V-α2.

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-ß (transforming growth factor-ß), 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.

Effects of single bouts of different endurance exercises with different intensities on microRNA biomarkers with and without blood flow restriction: a three-arm, randomized crossover trial.

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 < 0.001). The high-intensity intervention (HI) resulted in a higher LC than both lower intensity protocols (p < 0.001). The low-intensity blood flow restriction (LI-BFR) protocol led to a higher HR and higher LC than the low-intensity (LI) protocol without BFR (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 < 0.05). After LI exercise, the expression of miR-342-3p was up-regulated (p = 0.038). In LI-BFR, the level of miR-342-3p and miR-424-5p was confirmed to be up-regulated (p < 0.05). Three miRNAs and LC show a significant negative correlation (miR-99a-5p, 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.

Purinergic Regulation of Endothelial Barrier Function.

Increased vascular permeability is a hallmark of several cardiovascular anomalies, including ischaemia/reperfusion injury and inflammation. During both ischaemia/reperfusion and inflammation, massive amounts of various nucleotides, particularly adenosine 5'-triphosphate (ATP) and adenosine, are released that can induce a plethora of signalling pathways via activation of several purinergic receptors and may affect endothelial barrier properties. The nature of the effects on endothelial barrier function may depend on the prevalence and type of purinergic receptors activated in a particular tissue. In this review, we discuss the influence of the activation of various purinergic receptors and downstream signalling pathways on vascular permeability during pathological conditions.

SPARCL1 as a biomarker of maladaptive right ventricular remodelling in pulmonary hypertension.

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 < 0.01), DCM (p < 0.001) or controls (p < 0.001). Patients with adaptive RV function had higher SPARCL1 concentrations than controls (p < 0.05), whereas there was no difference between adaptive RV and DCM. SPARCL1 showed good predictive power for maladaptive RV (AUC 0.77, p < 0.001) with an optimal cut-off value of 9.66 ng/ml. The TAPSE/PASP ratio was the only independent predictor of SPARCL1 ≥ 9.66 ng/ml in multivariable logistic regression analysis.Conclusion: SPARCL1 shows potential as novel biomarker of RV pathological remodelling and is associated with RV maladaptation and ventriculoarterial uncoupling in PH.

IL10 Alters Peri-Collateral Macrophage Polarization and Hind-Limb Reperfusion in Mice after Femoral Artery Ligation.

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-/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.

Inhibition of Protein Prenylation of GTPases Alters Endothelial Barrier Function.

The members of Rho family of GTPases, RhoA and Rac1 regulate endothelial cytoskeleton dynamics and hence barrier integrity. The spatial activities of these GTPases are regulated by post-translational prenylation. In the present study, we investigated the effect of prenylation inhibition on the endothelial cytoskeleton and barrier properties. The study was carried out in human umbilical vein endothelial cells (HUVEC) and protein prenylation is manipulated with various pharmacological inhibitors. Inhibition of either complete prenylation using statins or specifically geranylgeranylation but not farnesylation has a biphasic effect on HUVEC cytoskeleton and permeability. Short-term treatment inhibits the spatial activity of RhoA/Rho kinase (Rock) to actin cytoskeleton resulting in adherens junctions (AJ) stabilization and ameliorates thrombin-induced barrier disruption whereas long-term inhibition results in collapse of endothelial cytoskeleton leading to increased basal permeability. These effects are reversed by supplementing the cells with geranylgeranyl but not farnesyl pyrophosphate. Moreover, long-term inhibition of protein prenylation results in basal hyper activation of RhoA/Rock signaling that is antagonized by a specific Rock inhibitor or an activation of cAMP signaling. In conclusion, inhibition of geranylgeranylation in endothelial cells (ECs) exerts biphasic effect on endothelial barrier properties. Short-term inhibition stabilizes AJs and hence barrier function whereas long-term treatment results in disruption of barrier properties.

Effects on the Profile of Circulating miRNAs after Single Bouts of Resistance Training with and without Blood Flow Restriction-A Three-Arm, Randomized Crossover Trial.

BACKGROUND: The effects of blood flow restriction (training) may serve as a model of peripheral artery disease. In both conditions, circulating micro RNAs (miRNAs) are suggested to play a crucial role during exercise-induced arteriogenesis. We aimed to determine whether the profile of circulating miRNAs is altered after acute resistance training during blood flow restriction (BFR) as compared with unrestricted low- and high-volume training, and we hypothesized that miRNA that are relevant for arteriogenesis are affected after resistance training. METHODS: Eighteen healthy volunteers (aged 25 ± 2 years) were enrolled in this three-arm, randomized-balanced crossover study. The arms were single bouts of leg flexion/extension resistance training at (1) 70% of the individual single-repetition maximum (1RM), (2) at 30% of the 1RM, and (3) at 30% of the 1RM with BFR (artificially applied by a cuff at 300 mm Hg). Before the first exercise intervention, the individual 1RM (N) and the blood flow velocity (m/s) used to validate the BFR application were determined. During each training intervention, load-associated outcomes (fatigue, heart rate, and exhaustion) were monitored. Acute effects (circulating miRNAs, lactate) were determined using pre-and post-intervention measurements. RESULTS: All training interventions increased lactate concentration and heart rate (p < 0.001). The high-intensity intervention (HI) resulted in a higher lactate concentration than both lower-intensity training protocols with BFR (LI-BFR) and without (LI) (LI, p = 0.003; 30% LI-BFR, p = 0.008). The level of miR-143-3p was down-regulated by LI-BFR, and miR-139-5p, miR-143-3p, miR-195-5p, miR-197-3p, miR-30a-5p, and miR-10b-5p were up-regulated after HI. The lactate concentration and miR-143-3p expression showed a significant positive linear correlation (p = 0.009, r = 0.52). A partial correlation (intervention partialized) showed a systematic impact of the type of training (LI-BFR vs. HI) on the association (r = 0.35 remaining after partialization of training type). CONCLUSIONS: The strong effects of LI-BFR and HI on lactate- and arteriogenesis-associated miRNA-143-3p in young and healthy athletes are consistent with an important role of this particular miRNA in metabolic processes during (here) artificial blood flow restriction. BFR may be able to mimic the occlusion of a larger artery which leads to increased collateral flow, and it may therefore serve as an external stimulus of arteriogenesis.

Targeted Ablation of Periostin-Expressing Activated Fibroblasts Prevents Adverse Cardiac Remodeling in Mice.

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.

Cardiac inflammation in genetic dilated cardiomyopathy caused by MYBPC3 mutation.

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 3months 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±0.4mg/mm, 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 (1.3±0.1% vs. 2.4±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±0.5pg/mL, p=0.02) in interleukin (IL)-6 compared to WT animals. Furthermore, RNA-seq 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.

Release kinetics of inflammatory biomarkers in a clinical model of acute myocardial infarction.

RATIONALE: Inflammation in the setting of acute myocardial infarction (MI) has been linked to risk stratification; however, the release kinetics of inflammatory biomarkers in patients with acute MI has been difficult to establish. OBJECTIVE: The aim of this study was to determine the kinetics of changes in the levels of several biomarkers specifically linked to inflammation after transcoronary ablation of septal hypertrophy, a procedure that mimics acute MI. METHODS AND RESULTS: We analyzed release kinetics of C-reactive protein, high-sensitivity C-reactive protein, interleukin-6, soluble CD40 ligand, and peripheral blood leukocyte subsets in patients (n=21) undergoing transcoronary ablation of septal hypertrophy. Blood samples were collected before transcoronary ablation of septal hypertrophy and at various times after transcoronary ablation of septal hypertrophy. Serum levels of C-reactive protein were increased at 24 hours (1.0 mg/dL [interquartile range [IQR], 0.7-1.75] versus 0.2 mg/dL [IQR, 0.1-1.05] at baseline [BL]; P<0.001), whereas high-sensitivity C-reactive protein increased as early as 8 hours (2.68 mg/L [IQR, 1.23-11.80] versus 2.17 mg/L [IQR, 1.15-5.06] at BL; P=0.002). Interleukin-6 was significantly increased at 45 minutes (2.59 pg/mL [IQR, 1.69-5.0] versus 1.5 pg/mL [IQR, 1.5-2.21] at BL; P=0.002), and soluble CD40 ligand was significantly decreased at 60 minutes (801.6 pg/mL [IQR, 675.0-1653.5] versus 1750.0 pg/mL [IQR, 1151.0-2783.0] at BL; P=0.016). Elevated counts of polymorphonuclear neutrophils were detectable at 15 minutes, with a significant increase at 2 hours (6415 cells/µL [IQR, 5288-7827] versus 4697 cells/µL [IQR, 2892-5620] at BL; P=0.004). Significant monocytosis was observed at 24 hours (729 cells/µL [IQR, 584-1344] versus 523 cells/µL [IQR, 369-701] at BL; P=0.015). CONCLUSIONS: Interleukin-6 and neutrophil granulocytes showed a continuous rise at all prespecified time points after induction of MI. Our results provide valuable additional evidence of the diagnostic value of inflammatory biomarkers in the setting of early acute MI.

Osteoglycin prevents cardiac dilatation and dysfunction after myocardial infarction through infarct collagen strengthening.

RATIONALE: To maintain cardiac mechanical and structural integrity after an ischemic insult, profound alterations occur within the extracellular matrix. Osteoglycin is a small leucine-rich proteoglycan previously described as a marker of cardiac hypertrophy. OBJECTIVE: To establish whether osteoglycin may play a role in cardiac integrity and function after myocardial infarction (MI). METHODS AND RESULTS: Osteoglycin expression is associated with collagen deposition and scar formation in mouse and human MI. Absence of osteoglycin in mice resulted in significantly increased rupture-related mortality with tissue disruption, intramyocardial bleeding, and increased cardiac dysfunction, despite equal infarct sizes. Surviving osteoglycin null mice had greater infarct expansion in comparison with wild-type mice because of impaired collagen fibrillogenesis and maturation in the infarcts as revealed by electron microscopy and collagen polarization. Absence of osteoglycin did not affect cardiomyocyte hypertrophy in the remodeling remote myocardium. In cultured fibroblasts, osteoglycin knockdown or supplementation did not alter transforming growth factor-ß signaling. Adenoviral overexpression of osteoglycin in wild-type mice significantly improved collagen quality, thereby blunting cardiac dilatation and dysfunction after MI. In osteoglycin null mice, adenoviral overexpression of osteoglycin was unable to prevent rupture-related mortality because of insufficiently restoring osteoglycin protein levels in the heart. Finally, circulating osteoglycin levels in patients with heart failure were significantly increased in the patients with a previous history of MI compared with those with nonischemic heart failure and correlated with survival, left ventricular volumes, and other markers of fibrosis. CONCLUSIONS: Increased osteoglycin expression in the infarct scar promotes proper collagen maturation and protects against cardiac disruption and adverse remodeling after MI. In human heart failure, osteoglycin is a promising biomarker for ischemic heart failure.

Release kinetics of high-sensitivity cardiac troponins I and T and troponin T upstream open reading frame peptide (TnTuORF) in clinically induced acute myocardial infarction.

CONTEXT: Troponin T upstream open reading frame peptide (TnTuORF) may be useful as a novel biomarker in acute cardiac syndromes. OBJECTIVE: The study examined the early release kinetics of TnTuORF. MATERIALS AND METHODS: We analyzed the time course of the release of cardiac troponins I and T and TnTuORF in patients (n = 31) with hypertrophic obstructive cardiomyopathy undergoing transcoronary ablation of septal hypertrophy (TASH). RESULTS: Fifteen minutes after TASH, the levels of both troponins increased significantly (cTnT median: 18 ng/L versus 27 ng/L; cTnI median: 15 ng/L versus 25 ng/L). TnTuORF showed no variation. DISCUSSION: We observed a significantly greater increase in cTnI compared with cTnT. CONCLUSION: Our results demonstrate that troponin assays allow early detection of myocardial injury, whereas TnTuORF levels remain unchanged in this setting.

Uridine Triphosphate Thio Analogues Inhibit Platelet P2Y12 Receptor and Aggregation.

Platelet P2Y12 is an important adenosine diphosphate (ADP) receptor that is involved in agonist-induced platelet aggregation and is a valuable target for the development of anti-platelet drugs. Here we characterise the effects of thio analogues of uridine triphosphate (UTP) on ADP-induced platelet aggregation. Using human platelet-rich plasma, we demonstrate that UTP inhibits P2Y12 but not P2Y1 receptors and antagonises 10 µM ADP-induced platelet aggregation in a concentration-dependent manner with an IC50 value of ~250 °µM. An eight-fold higher platelet inhibitory activity was observed with a 2-thio analogue of UTP (2S-UTP), with an IC50 of 30 µM. The 4-thio analogue (4S-UTP) with an IC50 of 7.5 µM was 33-fold more effective. A three-fold decrease in inhibitory activity, however, was observed by introducing an isobutyl group at the 4S- position. A complete loss of inhibition was observed with thio-modification of the γ phosphate of the sugar moiety, which yields an enzymatically stable analogue. The interaction of UTP analogues with P2Y12 receptor was verified by P2Y12 receptor binding and cyclic AMP (cAMP) assays. These novel data demonstrate for the first time that 2- and 4-thio analogues of UTP are potent P2Y12 receptor antagonists that may be useful for therapeutic intervention.

N-terminal fragment of cardiac myosin binding protein-C triggers pro-inflammatory responses in vitro.

Myocardial infarction (MI) leads to loss and degradation of contractile cardiac tissue followed by sterile inflammation of the myocardium through activation and recruitment of innate and adaptive cells of the immune system. Recently, it was shown that cardiac myosin binding protein-C (cMyBP-C), a protein of the cardiac sarcomere, is degraded following MI, releasing a predominant N-terminal 40-kDa fragment (C0C1f) into myocardial tissue and the systemic circulation. We hypothesized that early release of C0C1f contributes to the initiation of inflammation and plays a key role in recruitment and activation of immune cells. Therefore, we investigated the role of C0C1f on macrophage/monocyte activation using both mouse bone marrow-derived macrophages and human monocytes. Here we demonstrate that C0C1f leads to macrophage/monocyte activation in vitro. Furthermore, C0C1f induces strong upregulation of pro-inflammatory cytokines (interleukin-6 (IL-6), tumor necrosis factor α (TNFα), and interleukin-1ß (IL-1ß)) in cultured murine macrophages and human monocytes, resulting in a pro-inflammatory phenotype. We identified the toll-like receptor 4 (TLR4), toll-like receptor 2 (TLR2), and Advanced Glycosylation End Product-Specific Receptor (RAGE) as potential receptors for C0C1f whose activation leads to mobilization of the NFκB signaling pathway, a central mediator of the pro-inflammatory signaling cascade. Thus, C0C1f appears to be a key player in the initiation of inflammatory processes and might also play an important role upon MI.

Long-term verification of functional and structural renal damage after renal sympathetic denervation.

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 < 45 mL/min) during the early post- procedural period or the long-term follow-up (P = 0.34). The results of the present study show a sustained effect of RSD on BP reduction after a three-year follow-up, and there was no evidence of renal failure. These results provide verification of the long-term safety and effectiveness of RSD, even in patients with impaired renal function. © 2015 Wiley Periodicals, Inc.

Plasma microRNA-21 for the early prediction of acute kidney injury in patients undergoing major cardiac surgery.

BACKGROUND: Acute kidney injury (AKI) is a complication after major cardiac surgery that is associated with higher rates of morbidity and mortality. MicroRNA-21 (miR-21) has been described as an early biomarker for AKI. We investigated whether miR-21 is predictive of AKI and long-term mortality after cardiac surgery. METHODS: Consecutive patients (n = 115) undergoing major cardiac surgery were included. Serum creatinine was measured prior to, 4 h after, and 1, 4 and 7 days after extracorporeal circulation. Diagnosis of post-operative AKI was made in accordance with the international Kidney Disease: Improving Global Outcomes definition of AKI. Serum cystatin C and miR-21 were measured prior to and 4 h after surgery. miR-21 was determined by quantitative RT-PCR and was normalized to miRNA-39 from Caenorhabditis elegans. The median follow-up time was 2.9 years. RESULTS: AKI occurred in 36.5% (n = 42) of all patients. Baseline miR-21 was significantly lower in patients developing cardiac surgery-associated AKI (CSA-AKI) than in patients without CSA-AKI [0.27 (interquartile range, IQR, 0.14-0.30) versus 0.44 (IQR 0.25-0.75); P < 0.01]. Baseline miR-21 predicted CSA-AKI Stage 2/3 with an area under the curve of 0.701 [95% confidence interval (CI) 0.59-0.82; P = 0.007]. Baseline miR-21 <0.31 showed a hazard ratio of 3.11 (95% CI: 1.33-11.26) for CSA-AKI Stage 2/3. Patients with AKI Stage 2/3 had a significantly higher mortality (50 versus 10%; P = 0.0001) and dialysis rate (27 versus 11%; P = 0.038) within the 2.9-year follow-up. CONCLUSIONS: Our results indicate that miR-21 has the potential to identify patients at higher risk for CSA-AKI. This predictive value might be helpful in pre-procedural risk assessment and peri-procedural diagnosis and treatment.