Recurrence After Atrial Fibrillation Ablation and Investigational Biomarkers of Cardiac Remodeling.

BACKGROUND: Recurrence after atrial fibrillation (AF) ablation remains common. We evaluated the association between recurrence and levels of biomarkers of cardiac remodeling, and their ability to improve recurrence prediction when added to a clinical prediction model. METHODS AND RESULTS: Blood samples collected before de novo catheter ablation were analyzed. Levels of bone morphogenetic protein-10, angiopoietin-2, fibroblast growth factor-23, insulin-like growth factor-binding protein-7, myosin-binding protein C3, growth differentiation factor-15, interleukin-6, N-terminal pro-brain natriuretic peptide, and high-sensitivity troponin T were measured. Recurrence was defined as ≥30 seconds of an atrial arrhythmia 3 to 12 months postablation. Multivariable logistic regression was performed using biomarker levels along with clinical covariates: APPLE score (Age >65 years, Persistent AF, imPaired eGFR [<60 ml/min/1.73m2], LA diameter ≥43 mm, EF <50%; which includes age, left atrial diameter, left ventricular ejection fraction, persistent atrial fibrillation, and estimated glomerular filtration rate), preablation rhythm, sex, height, body mass index, presence of an implanted continuous monitor, year of ablation, and additional linear ablation. A total of 1873 participants were included. A multivariable logistic regression showed an association between recurrence and levels of angiopoietin-2 (odds ratio, 1.08 [95% CI, 1.02-1.15], P=0.007) and interleukin-6 (odds ratio, 1.02 [95% CI, 1.003-1.03]; P=0.02). The area under the receiver operating characteristic curve of a model that only contained clinical predictors was 0.711. The addition of any of the 9 studied biomarkers to the predictive model did not result in a statistically significant improvement in the area under the receiver operating characteristic curve. CONCLUSIONS: Higher angiopoietin-2 and interleukin-6 levels were associated with recurrence after atrial fibrillation ablation in multivariable modeling. However, the addition of biomarkers to a clinical prediction model did not significantly improve recurrence prediction.

Plasma angiopoietin-2 and its association with heart failure in patients with atrial fibrillation.

AIMS: Several biomarkers are associated with clinical outcomes in patients with atrial fibrillation (AF), but a causal relationship has not been established. This study aimed to evaluate angiopoietin-2, a novel candidate biomarker of endothelial inflammation and vascular remodelling, in patients with AF. METHODS AND RESULTS: Angiopoietin-2 was measured in plasma obtained from patients with AF treated with aspirin monotherapy (exploration cohort, n = 2987) or with oral anticoagulation (validation cohort, n = 13 079). Regression models were built to assess the associations between angiopoietin-2, clinical characteristics, and outcomes. In both cohorts, plasma angiopoietin-2 was independently associated with AF on the baseline electrocardiogram and persistent/permanent AF, age, history of heart failure, female sex, tobacco use/smoking, body mass index, renal dysfunction, diabetes, and N-terminal pro-B-type natriuretic peptide (NT-proBNP). Angiopoietin-2 was independently associated with subsequent hospitalization for heart failure after adjusting for age, creatinine, and clinical characteristics in the exploration cohort [c-index 0.79, 95% confidence interval (CI) 0.75-0.82; third vs. first quartile, hazard ratio (HR) 1.74, 95% CI 1.26-2.41] and in the validation cohort (c-index 0.76, 95% CI 0.74-0.78; HR 1.58, 95% CI 1.37-1.82). In both cohorts, the association persisted when also adjusting for NT-proBNP (P ≤ 0.001). In full multivariable models also adjusted for NT-proBNP, angiopoietin-2 did not show statistically significant associations with ischaemic stroke, cardiovascular and all-cause death, or major bleeding that were consistent across the two cohorts. CONCLUSIONS: In patients with AF, plasma levels of angiopoietin-2 were independently associated with subsequent hospitalization for heart failure and provided incremental prognostic value to clinical risk factors and NT-proBNP.

A retrospective study on poult enteritis syndrome in Minnesota.

A retrospective study was conducted to determine the occurrence of poult enteritis syndrome (PES) in Minnesota from January 2002 to December 2007. PES is an infectious intestinal disease of young turkeys between 1 day and 7 wk of age and is characterized by diarrhea, depression, and lethargy with pale intestines and/or excessively fluid cecal contents. During the study period, samples from 1736 turkey flocks were submitted to the Minnesota Veterinary Diagnostic Laboratory for disease investigation. Of these, 151 flocks (8.7%) were PES positive. Cases of PES were seen throughout the year with higher prevalence in fall. The PES was statistically associated with age with higher occurrence in poults less than 3 wk of age. Rotavirus, small round virus (SRV), Salmonella, nonhemolytic Escherichia coli, Enterococcus, and Eimeria oocysts were detected alone or in different combinations. Reovirus and adenovirus were found in one flock each. The most commonly identified pathogens were Salmonella (85 flocks) and rotavirus (73 flocks). Of PES-affected flocks, 39 (25.8%), 66 (43.7%), and 37 (24.5%) had one, two, and three or more pathogens, respectively. Rotavirus, SRV, and reovirus occurred mostly in poults of less than 6 wk of age while Salmonella, E. coli, and Eimeria were seen in poults of all age groups. Minimum age for rotavirus detection was in 2-day-old poults. Histopathologically, moderate to severe mixed intestinal villus or lamina propria inflammatory infiltrates, necrosis of distal villus tips in intestinal specimens, and mild to severe lymphocellular depletion in thymus, bursa, and spleen were seen. Antimicrobial sensitivity patterns of bacterial isolates from PES-affected flocks revealed maximum sensitivity to trimethoprim/sulfamethoxazole and ceftiofur and a varying degree of resistance to other antimicrobials.

Thermodynamic studies and binding mechanisms of cell-penetrating peptides with lipids and glycosaminoglycans.

Cell-penetrating peptides (CPPs) traverse the membrane of biological cells at low micromolar concentrations and are able to take various cargo molecules along with. Despite large differences in their chemical structure, CPPs share the structural similarity of a high cationic charge density. This property confers to them the ability to bind electrostatically membrane constituents such as anionic lipids and glycosaminoglycans (GAGs). Controversies exist, however, about the biological response after the interaction of CPPs with such membrane constituents. Present review compares thermodynamic binding studies with conditions of the biological CPP uptake. It becomes evident that CPPs enter biological cells by different and probably competing mechanisms. For example, some amphipathic CPPs traverse pure lipid model membranes at low micromolar concentrations--at least in the absence of cargos. In contrast, no direct translocation at these conditions is observed for non-amphipathic CPPs. Finally, CPPs bind GAGs at low micromolar concentrations with potential consequences for endocytotic pathways.

Binding and clustering of glycosaminoglycans: a common property of mono- and multivalent cell-penetrating compounds.

Recent observations in cell culture provide evidence that negatively charged glycosaminoglycans (GAGs) at the surface of biological cells bind cationic cell-penetrating compounds (CPCs) and cluster during CPC binding, thereby contributing to their endocytotic uptake. The GAG binding and clustering occur in the low-micromolar concentration range and suggest a tight interaction between GAGs and CPCs, although the relation between binding affinity and specificity of this interaction remains to be investigated. We therefore measured the GAG binding and clustering of various mono- and multivalent CPCs such as DNA transfection vectors (polyethylenimine; 1,2-dioleoyl-3-trimethylammonium-propane), amino acid homopolymers (oligoarginine; oligolysine), and cell-penetrating peptides (Penetratin; HIV-1 Tat) by means of isothermal titration calorimetry and dynamic light scattering. We find that these structurally diverse CPCs share the property of GAG binding and clustering. The binding is very tight (microscopic dissociation constants between 0.34 and 1.34 microM) and thus biologically relevant. The hydrodynamic radius of the resulting aggregates ranges from 78 nm to 586 nm, suggesting that they consist of numerous GAG chains cross-linked by CPCs. Likewise, the membrane-permeant monovalent cation acridine orange leads to GAG binding and clustering, in contrast to its membrane-impermeant structural analogs propidium iodide and ethidium bromide. Because the binding and clustering of GAGs were found to be a common denominator of all CPCs tested, these properties might be helpful to identify further CPCs.

High affinity of the cell-penetrating peptide HIV-1 Tat-PTD for DNA.

During cellular uptake of fluorescently labeled cell-penetrating peptides (CPPs), intense fluorescent signals are commonly observed in the nucleus of the cell, suggesting intracellular CPP relocation and potential binding to the genome of the host. We therefore investigated the interaction of the CPP HIV-1 Tat(47-57) with double-stranded DNA, and we also tested whether the fluorescence intensity of the labeled CPP allows for linear predictions of its intracellular concentration. Using isothermal titration calorimetry, we observe that the CPP has a high affinity for salmon sperm DNA as characterized by a microscopic dissociation constant of 126 nM. The binding is exothermic, with a reaction enthalpy of -4.63 kcal/mol CPP (28 degrees C). The dissociation constant and reaction enthalpy decrease further at higher temperatures. The affinity of the CPP for DNA is thus 1-2 magnitudes higher than for extracellular heparan sulfate, the likely mediator of the CPP uptake. Accordingly, the high affinity for DNA confers stability to extracellular transport complexes of CPP and DNA but potentially affects the regulation and molecular organization of the host's genome after nuclear uptake. Moreover, the CPP leads to the condensation of DNA as evidenced by the pronounced increase in light-scattering intensity. The fluorescence quantum yield of the FITC-labeled CPP decreases considerably at concentrations > 5 micromol/L, at pH < 7, and upon binding to DNA and glycosaminoglycans. This change in fluorescence quantum yield impedes the microscopic identification of uptake routes and the comparison of uptake efficiency of different CPPs, especially if the accumulation in subcellular compartments (self-quenching and pH difference) and transitory binding partners (quenching and condensation) is unknown.

Specific binding of Ro 09-0198 (cinnamycin) to phosphatidylethanolamine: a thermodynamic analysis.

Ro 09-0198 (cinnamycin) is a tetracyclic peptide antibiotic that is used to monitor the transbilayer movement of phosphatidylethanolamine (PE) in biological membranes during cell division and apoptosis. The molecule is one of the very rare examples where a small peptide binds specifically to a particular lipid. In model membranes and biological membranes containing phosphatidylethanolamine, Ro 09-0198 forms a 1:1 complex with this lipid. We have measured the thermodynamic parameters of complex formation with high sensitivity isothermal titration calorimetry and have investigated the structural consequences with deuterium and phosphorus solid-state NMR. Complex formation is characterized by a large binding constant, K0, of 10(7) to 10(8) M(-1), depending on the experimental conditions. The reaction enthalpy, DeltaHdegrees, varies between zero at 10 degrees C to strongly exothermic -10 kcal/mol at 50 degrees C. For large vesicles with a diameter of approximately 100 nm, DeltaHdegrees decreases linearly with temperature and the molar heat capacity of complex formation can be evaluated as = -245 cal/mol, indicating a hydrophobic binding mechanism. The free energy of binding is DeltaGdegrees = -10.5 kcal/mol and shows only little temperature dependence. The constancy of DeltaGdegrees together with the distinct temperature-dependence of DeltaHdegrees provide evidence for an entropy-enthalpy compensation mechanism: at 10 degrees C, complex formation is completely entropy-driven, at 50 degrees C it is enthalpy-driven. Varying the PE fatty acid chain-length between 6 and 18 carbon atoms produces similar binding constants and DeltaHdegrees values. Addition of Ro 09-0198 to PE containing bilayers eliminates the typical bilayer structure and produces 2H- and 31P-NMR spectra characteristic of slow isotropic tumbling. This reorganization of the lipid matrix is not limited to PE but also includes other lipids.