The presence of a large patent foramen ovale reduces acute and chronic success in atrial fibrillation ablation.

PURPOSE: In this study, we analyzed PFO implications in atrial fibrillation (AF) ablation. METHODS: Six hundred and twenty-five consecutive patients with AF undergoing PV isolation were included. We considered that a large and/or compliant PFO was present if the catheters advanced gently into the LA without puncturing the septum. Atrial tachyarrhythmias after the 3-month blanking period were classified as a recurrence. RESULTS: Out of the 625 patients included, 36 (5.8%) were found to have PFO. No significant differences were observed in the clinical characteristics of patients with PFO compared with patients without PFO. Nevertheless, patients with PFO had lower acute success in PV isolation compared with patients without PFO (98.2% vs. 88.5%; p = 0.006) even after adjusting for age, sex, type of AF, LA area, cardiomyopathy, time from AF diagnosis to the ablation, and ablation technique (odds ratio: 0.1; 95% confidence interval (CI): 0.02-0.9; p = 0.039). In 546 patients followed more than 6 months, the recurrence rate of any atrial tachyarrhythmia after 18.6 ± 11.9 months was significantly higher in patients with PFO compared with patients without PFO (41.9 vs. 70%; p = 0.012). This difference remained significant after adjusting for age, sex, type of AF, LA area, cardiomyopathy, time from AF diagnosis to the ablation, and ablation technique (hazard ratio: 1.9; 95% CI: 1.1-3.3; p = 0.015). CONCLUSIONS: The presence of a large and/or compliant PFO is an independent factor for PV isolation failure and arrhythmia recurrence rate after the ablation.

Polyaniline Nanofibers-Embedded Gold Nanoparticles Obtained by Template-Free Procedure with Immobilization Prospects.

In this work, template-free nanostructured conducting polymers (nCPs)-embedded gold nanoparticles (AuNPs) from aniline, thiophene and 3,4-ethylenedioxythiophene have been prepared via a one-pot sonochemical method. The synthesis of the nanocomposite (nCPs-AuNPs) was achieved in a short period of time (5-10 min), by applying high-energy ultrasound to an aqueous mixture of a CP precursor monomer and KAuCl4, in the presence of LiClO4 as dopant. The synthesis process is simpler, greener and faster in comparison to other procedures reported in the literature. Remarkably, bulk quantities of doped polyaniline PANI-AuNPs nanofibers were obtained. Subsequently, they were characterized by scanning electron microscopy (SEM), transmission electron microscopy (TEM), energy-dispersive X-ray spectroscopy (EDS) and Fourier transform infrared spectroscopy (FTIR), as well as by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). PANI-AuNPs nanofibers were also employed as immobilization matrix for a benchmark enzyme, glucose oxidase (GOX). Finally, glucose was determined in real samples of white and red wines by using the so-obtained GOX-PANI-AuNPs/Sonogel-Carbon biosensor, providing outstanding recoveries (99.54%). This work may offer important insights into the synthesis of nanostructured conducting polymers and also stimulates the exploration of the applications of these nanocomposites, especially in research fields such as (bio)sensors, catalysis and composite materials.

Simultaneous Detection of Glucose and Fructose in Synthetic Musts by Multivariate Analysis of Silica-Based Amperometric Sensor Signals.

Silica-based electrodes which permanently include a graphite/Au nanoparticles composite were tested for non-enzymatic detection of glucose and fructose. The composite material showed an effective electrocatalytic activity, to achieve the oxidation of the two analytes at quite low potential values and with good linearity. Reduced surface passivation was observed even in presence of organic species normally constituting real samples. Electrochemical responses were systematically recorded in cyclic voltammetry and differential pulse voltammetry by analysing 99 solutions containing glucose and fructose at different concentration values. The analysed samples consisted both in glucose and fructose aqueous solutions at pH 12 and in solutions of synthetic musts of red grapes, to test the feasibility of the approach in a real frame. Multivariate exploratory analyses of the electrochemical signals were performed using the Principal Component Analysis (PCA). This gave evidence of the effectiveness of the chemometric approach to study the electrochemical sensor responses. Thanks to PCA, it was possible to highlight the different contributions of glucose and fructose to the voltammetric signal, allowing their selective determination.

In Vitro Assay for Studying the Aggregation of Tau Protein and Drug Screening.

Aggregation of tau protein and formation of paired helical filaments is a hallmark of Alzheimer's disease and other tauopathies. Compared to other proteins associated with neurodegenerative diseases, the reported in vitro aggregation kinetics for tau protein are less consistent presenting a relatively high variability. Here we describe the development of an in vitro aggregation assay that mimics the expected steps associated with tau misfolding and aggregation in vivo. The assay uses the longest tau isoform (huTau441) which contains both N-terminal acidic inserts as well as four microtubule binding domains (MBD). The in vitro aggregation is triggered by addition of heparin and followed continuously by thioflavin T fluorescence in a 96 well microplate format. The tau aggregation assay is highly reproducible between different wells, experimental runs and batches of the protein. The aggregation leads to tau PHF-like morphology which is very efficient in seeding the formation of de novo fibrillar structures. In addition to its application in studying the mechanism of tau misfolding and aggregation, the current assay is a robust tool for screening drugs that could interfere with the pathogenesis of tau.

Cell-based Assay to Study Antibody-mediated Tau Clearance by Microglia.

Alzheimer's disease (AD) is a progressive neurodegenerative condition in which aggregated tau and amyloid proteins accumulate in the brain causing neuronal dysfunction which eventually leads to cognitive decline. Hyperphosphorylated tau aggregates in the neuron are believed to cause most of the pathology associated with AD. These aggregates are assumed to be released into the extracellular compartment and taken up by adjacent healthy neurons where they induce further tau aggregation. This "prion-like" spreading can be interrupted by antibodies capable of binding and "neutralizing" extracellular tau aggregates as shown in preclinical mouse models of AD. One of the proposed mechanisms by which therapeutic antibodies reduce pathology is antibody-mediated uptake and clearance of pathological aggregated forms of tau by microglia. Here, we describe a quantitative cell-based assay to assess tau uptake by microglia. This assay uses the mouse microglial cell line BV-2, allows for high specificity, low variability and medium throughput. Data generated with this assay can contribute to a better characterization of anti-tau antibody effector functions.

Enhancement of therapeutic potential of a naturally occurring human antibody targeting a phosphorylated Ser422 containing epitope on pathological tau.

Aggregation of tau protein and spreading of tau aggregates are pivotal pathological processes in a range of neurological disorders. Accumulating evidence suggests that immunotherapy targeting tau may be a viable therapeutic strategy. We have previously described the isolation of antibody CBTAU-22.1 from the memory B-cell repertoire of healthy human donors. CBTAU-22.1 was shown to specifically bind a disease-associated phosphorylated epitope in the C-terminus of tau (Ser422) and to be able to inhibit the spreading of pathological tau aggregates from P301S spinal cord lysates in vitro, albeit with limited potency. Using a combination of rational design and random mutagenesis we have derived a variant antibody with improved affinity while maintaining the specificity of the parental antibody. This affinity improved antibody showed greatly enhanced potency in a cell-based immunodepletion assay using paired helical filaments (PHFs) derived from human Alzheimer's disease (AD) brain tissue. Moreover, the affinity improved antibody limits the in vitro aggregation propensity of full length tau species specifically phosphorylated at position 422 produced by employing a native chemical ligation approach. Together, these results indicate that in addition to being able to inhibit the spreading of pathological tau aggregates, the matured antibody can potentially also interfere with the nucleation of tau which is believed to be the first step of the pathogenic process. Finally, the functionality in a P301L transgenic mice co-injection model highlights the therapeutic potential of human antibody dmCBTAU-22.1.

A common antigenic motif recognized by naturally occurring human VH5-51/VL4-1 anti-tau antibodies with distinct functionalities.

Misfolding and aggregation of tau protein are closely associated with the onset and progression of Alzheimer's Disease (AD). By interrogating IgG+ memory B cells from asymptomatic donors with tau peptides, we have identified two somatically mutated VH5-51/VL4-1 antibodies. One of these, CBTAU-27.1, binds to the aggregation motif in the R3 repeat domain and blocks the aggregation of tau into paired helical filaments (PHFs) by sequestering monomeric tau. The other, CBTAU-28.1, binds to the N-terminal insert region and inhibits the spreading of tau seeds and mediates the uptake of tau aggregates into microglia by binding PHFs. Crystal structures revealed that the combination of VH5-51 and VL4-1 recognizes a common Pro-Xn-Lys motif driven by germline-encoded hotspot interactions while the specificity and thereby functionality of the antibodies are defined by the CDR3 regions. Affinity improvement led to improvement in functionality, identifying their epitopes as new targets for therapy and prevention of AD.

Insoluble Off-Pathway Aggregates as Crowding Agents during Amyloid Fibril Formation.

The study of drug candidates for the treatment of amyloidosis and neurodegenerative diseases frequently involves in vitro measurements of amyloid fibril formation. Macromolecular crowding and off-pathway aggregation (OPA) are, by different reasons, two important phenomena affecting the scalability of amyloid inhibitors and their successful application in vivo. On the one hand, the cellular milieu is crowded with macromolecules that drastically increase the effective (thermodynamic) concentration of the amyloidogenic protein. On the other hand, off-pathway aggregates, rather than amyloid fibrils, are increasingly appointed as causative agents of toxicity. The present contribution reveals that insoluble off-pathway aggregates of hen egg-white lysozyme (HEWL) are a peculiar type of crowding agents that, unlike classical macromolecular crowders, decrease the thermodynamic concentration of protein. Illustrating this effect, OPA is shown to resume after lowering the fraction of insoluble aggregates at a constant soluble HEWL concentration. Protein depletion and thioflavin-T fluorescence progress curves indicate that OPA rebirth is not accompanied by additional amyloid fibril formation. The crystallization-like model extended to account for OPA and time-dependent activity coefficients is able to fit multiple kinetic results using a single set of three parameters describing amyloid nucleation, autocatalytic growth, and off-pathway nucleation. The list of fitted results notably includes the cases of aggregation rebirth and all types of progress curves measured for different HEWL concentrations. The quantitative challenges posed by macromolecular crowding and OPA find here a unified response with broader implications for the development of on- and off-pathway inhibitors.

Octreotide and enterocutaneous fistula closure in neonates and children.

UNLABELLED: Enterocutaneous fistula and its conservative management still pose a challenge for the surgeon. The use of octreotide and somatostatin in neonates and children as adjunctive therapy in the conservative management of this condition, leads to major controversy regarding its efficacy. Therefore, we conducted an extensive literature review of published articles regarding the use of somatostatin and its analogues in the treatment of enterocutaneous fistula in neonates and children. Our review is then presented together with a case vignette and discusses the different practical aspects of the treatment with these drugs. CONCLUSION: The major diversity in treatment regimens among published studies makes outcomes difficult to compare. However, given the results of the different cases reported in the literature and of our own experience, we suggest a possible beneficial effect of octreotide and somatostatin on closure of enterocutaneous fistula in these patients.

What Can the Kinetics of Amyloid Fibril Formation Tell about Off-pathway Aggregation?

Some of the most prevalent neurodegenerative diseases are characterized by the accumulation of amyloid fibrils in organs and tissues. Although the pathogenic role of these fibrils has not been completely established, increasing evidence suggests off-pathway aggregation as a source of toxic/detoxicating deposits that still remains to be targeted. The present work is a step toward the development of off-pathway modulators using the same amyloid-specific dyes as those conventionally employed to screen amyloid inhibitors. We identified a series of kinetic signatures revealing the quantitative importance of off-pathway aggregation relative to amyloid fibrillization; these include non-linear semilog plots of amyloid progress curves, highly variable end point signals, and half-life coordinates weakly influenced by concentration. Molecules that attenuate/intensify the magnitude of these signals are considered promising off-pathway inhibitors/promoters. An illustrative example shows that amyloid deposits of lysozyme are only the tip of an iceberg hiding a crowd of insoluble aggregates. Thoroughly validated using advanced microscopy techniques and complementary measurements of dynamic light scattering, CD, and soluble protein depletion, the new analytical tools are compatible with the high-throughput methods currently employed in drug discovery.

Enzyme kinetics: the whole picture reveals hidden meanings.

The methodology adopted by Michaelis and Menten in 1913 is still routinely used to characterize the catalytic power and selectivity of enzymes. These kinetic measurements must be performed soon after the purified enzyme is mixed with a large excess of substrate. Other time scales and solution compositions are no less physiologically relevant, but fall outside the range of applicability of the classical formalism. Here we show that the complete picture of an enzyme's mode of function is critically obscured by the limited scope of conventional kinetic analysis, even in the simplest case of a single active site without inhibition. This picture is now unveiled in a mathematically closed form that remains valid over the reaction time for all combinations of enzyme/substrate concentrations and rate constants. Algebraic simplicity is maintained in the new formalism when stationary reaction phases are considered. By achieving this century-old objective, the otherwise hidden role of the reversible binding step is revealed and atypical kinetic profiles are explained. Most singular kinetic behaviors are identified in a critical region of conditions that coincide with typical cell conditions. Because it is not covered by the Michaelis-Menten model, the critical region has been missed until now by low- and high-throughput screenings of new drugs. New possibilities are therefore raised for novel and once-promising inhibitors to therapeutically target enzymes.

Fluorinated beta-sheet breaker peptides.

The aggregation of amyloid-ß peptide (Aß) has been linked to the formation of neuritic plaques, which are pathological hallmarks of Alzheimer's disease. We synthesized peptides containing fluorinated amino acids and studied their effect on the Aß aggregation. The peptides were based on the sequence LVFFD, in which valine was substituted by either 4,4,4-trifluorovaline or 4-fluoroproline, or the phenylalanine at position 3 was replaced by 3,4,5-trifluorophenylalanine. Our results demonstrate that fluorination of the hydrophobic residue valine or phenylalanine is effective in preventing the Aß aggregation. This study opens up the possibility of using new sequences based on fluorinated amino acids to inhibit the amyloid-fibril formation.

A generic crystallization-like model that describes the kinetics of amyloid fibril formation.

Associated with neurodegenerative disorders such as Alzheimer, Parkinson, or prion diseases, the conversion of soluble proteins into amyloid fibrils remains poorly understood. Extensive "in vitro" measurements of protein aggregation kinetics have been reported, but no consensus mechanism has emerged until now. This contribution aims at overcoming this gap by proposing a theoretically consistent crystallization-like model (CLM) that is able to describe the classic types of amyloid fibrillization kinetics identified in our literature survey. Amyloid conversion represented as a function of time is shown to follow different curve shapes, ranging from sigmoidal to hyperbolic, according to the relative importance of the nucleation and growth steps. Using the CLM, apparently unrelated data are deconvoluted into generic mechanistic information integrating the combined influence of seeding, nucleation, growth, and fibril breakage events. It is notable that this complex assembly of interdependent events is ultimately reduced to a mathematically simple model, whose two parameters can be determined by little more than visual inspection. The good fitting results obtained for all cases confirm the CLM as a good approximation to the generalized underlying principle governing amyloid fibrillization. A perspective is presented on possible applications of the CLM during the development of new targets for amyloid disease therapeutics.

[Severe bilateral vesicoureteral reflux and nephropathy in male newborns: valve-like syndrome or uncoordinated voiding in the male fetus]. / Reflujo vesicoureteral bilateral grave y nefropatía renal en pacientes varones recién nacidos: síndrome de válvulas-like o Micción no coordinada en el feto varón.

OBJECTIVES: Various papers differentiating neonatal vesicoureteral reflux (VUR) with severe renal damage from other predominant group of newborns with neonatal VUR without renal lesions and those diagnosed in older ages, generally in relation with urinary tract infection (UTI), have been published over the last decade. From the standpoint that VUR is part of a broad spectrum both in clinical expression as in pathogenesis, with different theories described to explain the existence of this type of congenital VUR in males. The existence of a fetal vesicourethral dysfunction, presenting after birth as a high risk bladder, which is defined by urodynamic tests in the first trimester, explains the appearance of severe fetal VUR with functional deterioration of one or both renal units at the time of birth; this entity must be diagnosed to establish the adequate therapeutic management. This clinical picture is named Valve like syndrome or male uncoordinated fetal voiding.

[Current status of the retroperitoneoscopic surgical approach in pediatric urology]. / Estado actual del abordaje retroperitoneoscopico en urologia pediatrica.

Urologic pediatric retroperitoneoscopy has had three different stages that have conditioned, although not hindered, its development: 1) limited number of indications in the pediatric age, because pediatric surgery itself is not much invasive; 2) adaptation of the technological development from adult to children; 3) overcoming the controversies between laparoscopic and non laparoscopic pediatric surgeons. After overcoming these stages, retroperitoneoscopy has become an indispensable tool for the treatment of various diseases of the kidney, being nephrectomy the gold standard among the indications for ablation, and pyeloplasty among reconstructive, through a complete or assisted retroperitoneoscopic approach.

Smoking among pregnant women in Cantabria (Spain): trend and determinants of smoking cessation.

BACKGROUND: Cantabria (Spain) has one of the highest prevalence of smoking among women of the European Union. The objectives are to assess the trend of smoking during pregnancy in a five-year period and the determinants of smoking cessation during pregnancy in Cantabria. METHODS: A 1/6 random sample of all women delivering at the reference hospital of the region for the period 1998-2002 was drawn, 1559 women. Information was obtained from personal interview, clinical chart, and prenatal care records. In the analysis relative risks and 95% confidence intervals were estimated. Multivariable analysis was carried out using stepwise logistic regression. RESULTS: Smoking prior to pregnancy decreased from 53.6% in 1998 to 39.4% in 2002. A decrease in smoking cessation among women smoking at the beginning of pregnancy was observed, from 37.3% in 1998 to 20.6% in 2002. The mean number of cigarettes/day (cig/d) before pregnancy remained constant, around 16 cig/d, whereas a slight trend to increase over time was seen, from 7.7 to 8.9 cig/d. In univariate analysis two variables favoured significantly smoking cessation, although they were not included in the stepwise logistic regression analysis, a higher education level and to be married. The logistic regression model included five significant predictors (also significant in univariate analysis): intensity of smoking, number of previous pregnancies, partner's smoking status, calendar year of study period (these four variables favoured smoking continuation), and adequate prenatal care (which increased smoking cessation). CONCLUSION: The frequency of smoking among pregnant women is very high in Cantabria. As smoking cessation rate has decreased over time, a change in prenatal care programme on smoking counseling is needed. Several determinants of smoking cessation, such as smoking before pregnancy and partner's smoking, should be also addressed by community programmes.

[Treatment of rheumatoid arthritis with anakinra: a systematic review]. / Tratamiento de la artritis reumatoide con anakinra: revisión sistemática.

OBJECTIVE: To perform a systematic review for evaluating efficacy and safety of anakinra in the treatment of rheumatoid arthritis (RA). MATERIAL AND METHOD: The MedLine, Embase, and Cochrane Library databases were searched from January 2000 to February 2006 by using a high sensitive search that included every randomised controlled trial (RCTs) or controlled trial (CTs) that evaluated either efficacy or safety of Anakinra for the treatment of RA. RESULTS: The search identified four relevant studies to evaluate efficacy. Patients treated with anakinra achieved significantly better clinical responses than those treated with placebo. Anakinra combined with methotrexate provided significantly greater clinical benefit than methotrexate alone. Combination therapy with etanercept and anakinra provides no added benefit and an increased safety risk compared with etanercept alone. Results from a large, placebo-controlled safety study demonstrate that anakinra is safe and well tolerated. The most common adverse effect was a mild local inflammation over the puncture area. CONCLUSIONS: This review confirmed both the efficacy and the safety of anakinra in the short term for the treatment of RA. Anakinra provides adequate clinical responses without major safety problems. This systematic review does not allow us to conclude on Anakinra responses in the long term.

Weekday but not weekend alcohol consumption before pregnancy influences alcohol cessation during pregnancy.

BACKGROUND: Cantabria has the highest prevalence of alcohol consumption among women in Spain. Patterns of alcohol consumption before pregnancy were assessed as a determinant of alcohol cessation in pregnant women in Cantabria. METHODS: Survey on a random sample of women delivering for the period 1998-2002 (n = 1510). Information was obtained from personal interview (data on alcohol consumption), clinical charts and prenatal care records. Relative risks (RR) and 95% confidence intervals (CI) were estimated. Multivariable analyses were carried out using logistic regression. RESULTS: Nearly half (49.5%) of the women drank regularly before pregnancy and 22.7% during pregnancy. Sociodemographic variables favouring alcohol cessation were: high education level and smoking cessation, whereas high social class, advanced maternal age and employment outside of home decreased the rate of alcohol cessation. Cessation decreased with the amount of alcohol consumed on weekdays (P < 0.001), but not with intake during weekends only. In women with alcohol use only during weekends, only the consumption of spirits increased the rate of alcohol cessation (adjusted RR = 1.40, 95% CI: 1.13-1.60). Pre-pregnancy binge drinking (> or =4 drinks on one occasion) decreased alcohol cessation in pregnancy (adjusted RR = 0.66, 95% CI: 0.40-0.97). CONCLUSIONS: Drinking patterns influenced the rate of alcohol cessation: the heavier the alcohol consumption on weekdays, the lower the rate of alcohol cessation.

Pattern of alcohol consumption during pregnancy and risk for low birth weight.

PURPOSE: The effects of moderate alcohol drinking on low birth weight are not clear, and conflicting results have been reported. We assessed the influence of different patterns of alcohol drinking during pregnancy on low birth weight. METHODS: A case-control study was carried out at the University of Cantabria Hospital, Spain, from 1998 to 2002. Cases (n = 552) were mothers delivering a single newborn weighing less than 2500 g. Controls (n = 1451) were selected from a random sample of all delivering women. Information was obtained from personal interview, clinical charts, and prenatal care records. Odds ratios (ORs) and 95% confidence intervals (CIs) were estimated. Adjustment for potential confounders was made by means of logistic regression. RESULTS: Alcohol consumption of less than 6 g/day decreased the risk for low birth weight (adjusted OR = 0.64; 95% CI, 0.46-0.88). A similar result was obtained for moderate drinkers (<12 g/day) on weekends only. The opposite relationship was observed between alcohol consumption on weekdays of 12 g/day or greater (adjusted OR = 2.67; 95% CI, 1.39-5.12), not observed in those drinking on weekends only. The interaction between alcohol consumption and tobacco smoking was analyzed. Weekday drinkers of 12 g/day or greater showed an increased risk in smokers. Alcohol consumption on weekends only in nonsmokers was inversely related. The influence of alcohol was greater for small-for-gestational-age (SGA) than non-SGA babies. CONCLUSIONS: Alcohol consumption of 12 g/day or greater increased the risk for low birth weight, whereas lower consumption during weekends showed the opposite effect (mainly in nonsmokers).

Breast-feeding and risk of hospitalization for all causes and fever of unknown origin.

BACKGROUND: Breast-feeding has shown to give a reduction in the risk of hospitalization due to respiratory tract infections and gastrointestinal conditions during the first two years of life. The association of breast-feeding with all admission causes and with fever of unknown origin (FUO) was analysed. METHODS: A case-reference study in Cantabria (northern Spain) was carried out. Cases (n=336) were hospitalized children aged less than 24 months at University of Cantabria Hospital; the reference was a 1:1 matched (by time from delivery to admission) sample of children from mothers delivering at the same hospital. Information on breast-feeding, socioeconomic variables and employment were obtained. Odds ratios (ORs), their 95% confidence intervals (CIs), and mean length of breast-feeding were estimated after adjusment for confounding variables. RESULTS: In the reference population, shorter duration of breast-feeding was associated with smoking, lower educational level, and less privileged social strata. The frequency of breast-feeding was higher in the reference than in the cases, 82.3% vs 75.6% (p=0.023). Significant negative trends were noted in univariate analyses between the length of breast-feeding and both all admission causes and FUO, although the statistical significance was lost after adjusting for confounding variables (educational level, social class, smoking, and use of incubator after delivery). The adjusted mean length of breast-feeding was shorter in hospitalized children < or = 6 months old for both all admission causes (40.6 +/- 5.4 vs 99.5 +/- 5.4, p < 0.001) and FUO (40.8 +/- 12.4 vs 91.7 +/- 12.4, p=0.006). CONCLUSION: Breast-feeding time is shorter in hospitalized children for both all admission causes and FUO.