Same-day discharge after catheter ablation for routine arrhythmias: an initial experience.

AIM: Same-day discharge (SDD) in the setting of catheter ablation (CA) is not widely applied. We present our experience concerning SDD in a selected population of patients who underwent CA; the outcome was evaluated in terms of feasibility and safety. METHODS: 401 CA procedures were performed at our institution between January 2008 and December 2009 in 379 patients (65±16 years, 221 men). 336 CA procedures (84%) were considered eligible for SDD, after the exclusion of ventricular arrhythmias, atrial fibrillation, atypical atrial flutter, AV node ablation as well as procedures involving an arterial or transseptal access. Subsequently, a number of clinical and organizational exclusion criteria were applied. RESULTS: 223 patients were actually discharged on the same day of CA (56% of 401 overall CA procedures): 114 atrial flutter (AFL) and 109 supraventricular tachycardia. Many patients were excluded before CA due to a limited availability of the day-hospital facility; this occurred more frequently in the year 2008 than 2009 (45 vs. 2, P=0.0001); in the year 2009 the rate of total CA procedures which underwent SDD was of 68%. Overall, three groin hematomas occurred, all in patients ablated for AFL. Two of them were recognized during the postablation CONCLUSION: SDD can be safely performed in most patients undergoing CA for routine arrhythmias. This may result in a significant impact on daily practice in terms of both organizational improvement and subjective benefit for the patients.

Relationship between low levels of high-density lipoprotein cholesterol and dementia in the elderly. The InChianti study.

BACKGROUND: To evaluate the association between plasma lipid fractions and the prevalence of dementia in a large sample of Italian older individuals. METHODS: A total of 1051 older community-dwelling individuals (age >/=65 years), enrolled in the InChianti study, were included. Diagnosis of dementia was established at baseline and at the 3-year follow-up using Diagnostic and Statistical Manual of Mental Disorder (Fourth Edition) criteria. Plasma lipids were measured by standardized methods at baseline and after 3 years. RESULTS: At baseline, 61 individuals (5.8%) were affected by dementia. Demented individuals showed significantly lower total cholesterol (TC), nonhigh-density lipoprotein cholesterol, and high-density lipoprotein cholesterol (HDL-C) levels compared with controls; no differences were found in triglycerides (TG) and lipoprotein (a) levels. Of the 819 subjects reevaluated at the 3-year follow-up, 81 (9.9%) received a new diagnosis of dementia. Again, demented subjects were characterized by significantly lower TC, non-HDL-C, and HDL-C levels compared with controls, thus confirming the baseline findings. At multivariate logistic regression analysis, HDL-C levels (odds ratio: 0.96, 95% confidence interval: 0.93-0.99), but not TG and non-HDL-C, were associated with dementia independent of important confounders including age, gender, apo E phenotype, stroke, weight loss, interleukin 6 levels, and ankle-brachial index. CONCLUSIONS: Among community-dwelling older people, individuals affected by dementia showed significantly lower TC, non-HDL-C, and HDL-C levels; however, at multivariate analysis, only HDL-C was associated with dementia. Our results suggest the existence of an independent relationship between dementia and low HDL-C levels.

The role of polyunsaturated fatty acids (PUFA) in the treatment of dyslipidemias.

Polyunsaturated fatty acids (PUFA) are a family of lipids including some subgroups identified by the position of the last double bond in their structure. PUFA n-3 include alpha linolenic acid (ALA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA), while PUFA n-6 include linoleic acid (LA) and arachidonic acid (AA). Since PUFA n-3 consumption has been shown to be inversely correlated with coronary heart diseases (CHD) incidence, clinical trials have been principally conducted by administering fish oil supplements or purified PUFA n-3. The relationship between dietary PUFA n-3 and CHD is believed to be only partially mediated by their effects on plasma lipoprotein profile. PUFA n-3 have shown to reduce only slightly total and LDL cholesterol, probably as they crowd saturated fatty acids out of diet. Data on HDL cholesterol suggest that PUFA n-3 produce only a small increase in this fraction. The effect of PUFA n-3 supplementation on plasma triglycerides (TG) is much more important, with a reduction of about 25% in normolipidemic subjects and about 50% in hypertriglyceridemic patients. This effect seems to be mediated by an inhibition of hormone-sensitive lipase, and VLDL secretion, and an increase in apo B liver degradation. They also increase lipoprotein lipase activity resulting in a reduction of post-prandial TG. PUFA n-3 might be used as second line therapy, additional or alternative to fibrates and nicotinic acid, in the treatment of severe hypertriglyceridemia. Furthermore, the addition of PUFA n-3 to statin therapy might contribute to normalize TG levels in patients with combined hyperlipidemia.

Markers of endothelial dysfunction in older subjects with late onset Alzheimer's disease or vascular dementia.

A consistent amount of evidence suggests that vascular factors might be involved in the pathogenesis of late onset Alzheimer's disease (LOAD). We evaluated the presence of endothelial dysfunction by measuring the plasma levels of soluble E-selectin and vascular cell adhesion molecule 1 (VCAM-1) in a sample of patients affected by LOAD (n. 60) or vascular dementia (VD: n. 80). They were compared with a sample of older patients with cerebrovascular disease but not-dementia (CDND: n. 40), and with a sample of healthy older controls (n. 30). sVCAM-1 plasma levels were higher in LOAD and VD compared with controls. Among patients (LOAD, VD, and CDND), sE-selectin levels were higher in individuals with most severe cerebrovascular disease on CT scan. At multivariate regression analysis, fasting glucose (p<0.05) and TNF-alpha levels (p<0.02) were positively correlated with sE-selectin levels (adjusted r(2): 20%), while sVCAM-1 was positively correlated with age (p<0.01), and alcohol consumption (p: 0.03), and negatively associated with HDL-C levels (p: 0.005), (p<0.01; adjusted r(2): 44%), independent of possible confounders. Increased sVCAM-1 plasma levels in LOAD and VD suggest the existence of endothelial dysfunction in both types of dementia. The possible role of E-selectin in the pathogenesis of cerebrovascular disease is also supported by our data.

Monitoring 2-D gel-induced modifications of proteins by MALDI-TOF mass spectrometry.

In addition to more than 200 endogenously produced post-translational modifications, a detailed analysis of 2-D gel-separated proteins must also consider other modifications that a protein can experience during various steps of its separation. This review describes the use of matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry to investigate some of these modifications, which can originate during sample preparation and/or during the separation phase. The analyses described were mostly conducted at pH 9-9.5, and yielded reliable information on stable adduct formation that involved protein-bound amino acids and a number of gel components, including acrylamide derivatives, gel cross-linkers, and Immobiline chemicals. The -SH group of Cys was found to be the prime target of such adducts; however, longer reaction times revealed the involvement of the epsilon-NH2 of Lys. The same analysis revealed that the failure to achieve full reduction/alkylation prior to any electrophoretic step could result in protein-protein interaction, which could lead to a number of spurious spots in the final 2-D map. The implications of these modifications on the MS analysis in particular and on proteome research in general are discussed.

[Low-molecular weight heparin and acute coronary syndrome: theoretical background and its use in clinical practice]. / Eparine a basso peso molecolare e sindromi coronariche acute: razionale teorico ed uso nella pratica clinica.

Thrombosis is responsible for the acute manifestations of coronary artery disease. Intravenous heparin has been shown to be effective in reducing the risk of death or myocardial infarction in patients with acute coronary syndromes. Compared to standard heparin, low-molecular weight heparins (LMWHs) have improved pharmacological and pharmacokinetic properties. A number of LMWHs, such as nadroparin, dalteparin and enoxaparin, have been evaluated in patients with acute coronary syndromes. FRISC (Fragmin during Instability in Coronary Artery Disease) and FRIC (Fragmin in Unstable Coronary Artery Disease), evaluated dalteparin and found the LMWH to be more effective than aspirin alone (FRISC) and as effective as heparin in a direct comparison (FRIC). In a small trial, nadroparin was shown to significantly reduce the risk of ischemic outcomes compared with a combination of aspirin and heparin, but this effect was no longer significant in the large FRAX.I.S. trial (FRAXiparine in Ischaemic Syndrome). Enoxaparin resulted in a statistically significant reduction in the combined outcome of death, myocardial infarction and recurrent angina or of urgent revascularization when compared with heparin in the ESSENCE (Efficacy and Safety of Subcutaneous Enoxaparin in Non-Q-Wave Coronary Events) and TIMI 11B trials. Meta-analyzing of the data of these two trials revealed that even the combination of death and myocardial infarction was significantly reduced by the use of enoxaparin. There is accumulating evidence that LMWHs are safe and effective alternatives to standard heparin for the treatment of acute coronary syndromes and that they offer practical and therapeutic advantages.

Cardiac markers and risk stratification: an integrated approach.

Risk stratification of patients with acute coronary syndromes (ACS) is pivotal for correct allocation of health resources and for maximizing the benefit of available treatment modalities. However, clinical and electrocardiographic indicators of high risk lack sufficient sensitivity for the detection of major cardiac events. The complementary information provided by the measurement of different biomarkers is believed to be very useful. Specifically, elevations of cardiac troponin I (cTnI) and T (cTnT) are strongly associated with a high-risk profile both at short- and long-term. This has been definitely demonstrated in many studies as well as in cumulative meta-analysis. The role of different biomarkers, such as those reflecting activation of hemostasis and the presence of inflammation, is however less defined. At the moment, no study has prospectively evaluated these biomarkers in the whole spectrum of unselected patients with ACS. It is also unclear whether these biomarkers add independent prognostic value to the clinical and electrocardiographic indicators of adverse outcome and whether they offer additional information when compared to each other. The Early Prognostic Value of Biochemical Markers of Myocardial Damage, Activation of Hemostatic Mechanism and Inflammation in Acute Ischemic Syndromes (EMAI) study has been prospectively designed to solve these issues. In this study, we have evaluated the prognostic value of cTnI and cTnT, D-dimer, prothrombin fragment 1+2 (F1+2), thrombin-antithrombin complex (TAT) and C-reactive protein (CRP) in patients with ACS at the time of admission. We have enrolled in 31 Italian Coronary Care Units 1971 patients with rest anginal pain within 12 h from admission and electrocardiographic evidence of myocardial ischemia. Of these, 730 patients resulted to have ST-segment elevation myocardial infarction eligible for a reperfusion strategy and 1241, an acute coronary syndrome without persisting ST-segment elevation. Primary outcome measure of the study is the composite of death and non-fatal MI within 30 days from admission, which has occurred in 8.9% of the study population.

Prognostic role of hemostatic markers in acute coronary syndromes patients.

Coronary thrombosis is an important determinant of prognosis in patients with acute coronary syndromes (ACS). However, the identification of patients at high-risk for progression of coronary thrombosis is difficult in part because we currently lack clinically meaningful laboratory methods for its detection. The most promising approaches involve the measurement in plasma of markers of fibrin formation and degradation. Thrombin activity, as reflected by plasma or urine concentrations of fibrinopeptide A, is increased in patients with ACS and is associated with adverse outcome. However, the use of fibrinopeptide A as a marker of fibrin formation is limited by the very short half-life of the compound, by artifact due to sample acquisition, and by extremely long turnaround times. To overcome these limitations, measurement of soluble fibrin has been proposed. We have recently explored the prognostic value of a new fibrin-specific ELISA assay for soluble fibrin in patients with ACS and found that patients with highest levels had a twofold increased risk of early and late cardiac events. Increases in plasma concentrations of cross-linked fibrin degradation products (XL-FDPs), which reflect increased fibrin turn-over, are a marker of risk for complications of myocardial infarction. However, until recently, assays for XL-FDPs lacked specificity, because they did not distinguish between fibrin and fibrinogen degradation products. Recently, fibrin-specific ELISAs have been described and a rapid whole blood assay for D-dimer has been developed. We recently validated the prognostic value of this whole blood agglutination assay in patients with ACS. The results suggest that: (1) the detection of significant activation of the coagulation and/or fibrinolytic system may be important for rapid risk stratification of patients with ACS; (2) patients with biochemical evidence of ongoing coronary thrombosis may particularly benefit from aggressive antithrombotic strategies; (3) sequential measurements of these markers may be useful to guide antithrombotic treatment during the unstable phase of coronary artery disease.

Reduction and alkylation of proteins in preparation of two-dimensional map analysis: why, when, and how?

The standard procedure adopted up to the present in proteome analysis calls for just reduction prior to the isoelectric focusing/immobilized pH gradient (IEF/IPG) step, followed by a second reduction/alkylation step in between the first and second dimension, in preparation for the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) step. This protocol is far from being optimal. It is here demonstrated, by matrix assisted laser desorption/ionization-time of flight (MALDI-TOF)-mass spectrometry, that failure to reduce and alkylate proteins prior to any electrophoretic step (including the first dimension) results in a large number of spurious spots in the alkaline pH region, due to "scrambled" disulfide bridges among like and unlike chains. This series of artefactual spots comprises not only dimers, but an impressive series of oligomers (up to nonamers) in the case of simple polypeptides such as the human alpha- and beta-globin chains, which possess only one (alpha-) or two (beta-) -SH groups. As a result, misplaced spots are to be found in the resulting two-dimensional (2-D) map, if performed with the wrong protocol. The number of such artefactual spots can be impressively large. In the case of analysis of complex samples, such as human plasma, it is additionally shown that failure to alkylate proteins results in a substantial loss of spots in the alkaline gel region, possibly due to the fact that these proteins, at their pI, regenerate their disulfide bridges with concomitant formation of macroaggregates which become entangled with and trapped within the polyacrylamide gel fibers. This strongly quenches their transfer in the subsequent SDS-PAGE step.

Alkylation kinetics of proteins in preparation for two-dimensional maps: a matrix assisted laser desorption/ionization-mass spectrometry investigation.

All existing protocols for protein separation by two-dimensional (2-D) gel electrophoresis require the full reduction, denaturation, and alkylation as a precondition for an efficient and meaningful separation of such proteins. Existing literature provides a strong evidence to suggest that full reduction and denaturation can be achieved in a relatively short time; the same thing, however, can not be said for the alkylation process, which the present study shows that more than 6 h are required for a complete alkylation. We have used matrix assisted laser desorption/ionisation-time of flight-mass spectrometry (MALDI-TOF-MS) to monitor protein alkylation by iodoacetamide over the period 0-24 h at pH 9. The present, fast and specific MS method provided clear indication on the extent and speed of alkylation which reached approximately 70% in the first 2 min, yet the remaining 30% resisted complete alkylation up to 6 h. The use of sodium dodecyl sulfate (SDS) during the alkylation step resulted in a strong quenching of this reaction, whereas 2% 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) exerted a much reduced inhibition. The implications of the present measurements on 2-D gel analysis in particular and proteomics in general are discussed.

Protein alkylation in the presence/absence of thiourea in proteome analysis: a matrix assisted laser desorption/ionization-time of flight-mass spectrometry investigation.

Although it is highly recommended that reduction and alkylation of free -SH groups in proteins should be performed prior to any electrophoretic step (including the first isoelectric focusing/immobilized pH gradient (IEF/IPG) dimension), it is here reported that one component of the sample solubilization cocktail adopted recently (namely thiourea) strongly quenches such alkylation process (as typically carried out with iodoacetamide, IAA). The present matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS) analysis demonstrates that thiourea is an effective scavenger of IAA, since its sulfur atom reacts as efficiently as the ionized, free -SH group of Cys in proteins at alkaline pH values (pH 8.5-9.0). As a result of this reaction, free IAA is quickly depleted by thiourea, via the formation of an intermediate adduct, which is rapidly deamidated to form the cyclic compound thiazolinidone monoimine. This reaction strongly competes with the direct addition reaction of IAA onto the -SH group in proteins, resulting in poorly alkylated proteins. It is, therefore, recommended that, whenever possible and compatible with the type of sample, thiourea should be omitted from the solubilizing cocktail in proteome analysis. However, after proper sample reduction and alkylation, thiourea can be incorporated into the IEF/IPG gel, where it will have the beneficial effect of augmenting protein solubility at their pI values and scavenging the excess of free IAA.

Protein alkylation by acrylamide, its N-substituted derivatives and cross-linkers and its relevance to proteomics: a matrix assisted laser desorption/ionization-time of flight-mass spectrometry study.

The present review highlights some important alkylation pathways of proteins, as measured by matrix assisted laser desorption/ionization-time of flight (MALDI-TOF)-mass spectrometric analysis, engendered by acrylamide and a number of its derivatives, including N-substituted acrylamides, cross-linkers and Immobilines (the acrylamido weak acids and bases used to create immobilized pH gradients). The present data are of relevance in two-dimensional maps and proteome analysis. It is shown that acrylamide can alkylate the -SH group of proteins even when engaged in disulfide bridges. An order of reactivity is obtained for a series of cross-linkers, which are shown to have an extremely reacting double bond, with the second one almost unreactive, originating "pendant, unreacted ends", which can subtract proteins migrating in a gel by covalently affixing them to it. An analogous reactivity scale is constructed also for the Immobiline chemicals, whose reactivity is shown to be linearly dependent on the pK values, the least reacting species being the acidic compounds. When analyzing real-life samples by two-dimensional (2-D) maps, like milk powders, a number of modifications can be detected by MALDI-TOF mass spectra of eluted spots, including variable phosphorylation sites (up to nine) and lactosyl moieties. If, for eluting such spots, formic acid is used, MALDI-TOF mass spectrometry (MS) reveals an incredible number of formylation sites, on Ser and Thr residues.

Investigating the reaction of a novel silica capillary coating compound with proteins/peptides by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry.

Quaternized piperazine ((N-methyl-N-omega-iodobutyl-N'-methyl)piperazine; QPzl) is a novel compound described as an ideal coating material for the silica capillaries that are commonly used for capillary zone electrophoresis. In the course of such analysis, contact between such coatings and biomolecules may result in certain modifications of the latter. To gain specific information on such potential modifications, solutions at pH 10.0 containing both QPzl and standard proteins/peptides were incubated for various periods and examined by matrix-assisted laser desorption/ionisation mass spectrometry. The reduction of the S-S bridges, denaturation in 8 M urea, the isoelectric point of the protein and the duration of the incubation had a profound influence on the investigated reaction. Analysis in reflectron mode and post source decay identified Cys as the likely site of interaction. The implications of the present measurements for proteome analysis using capillary and gel electrophoresis are discussed.

Two-dimensional gel electrophoresis/matrix-assisted laser desorption/ionisation mass spectrometry of commercial bovine milk.

Proteins in commercial bovine milk have been separated by two-dimensional gel electrophoresis and examined by matrix-assisted laser desorption/ionisation mass spectrometry. Gel separation was conducted in two different pH gradients, 3-10 and 6-11; the latter range resulted in a higher spot resolution and favoured the basic proteins. We have limited the time-of-flight mass spectrometry analysis to the linear mode to examine the capability of reliable relative molecular masses of the intact proteins in their characterisation. The present study draws attention to the difficulty of identifying basic proteins with low molecular masses (below 12000 Da) that are commonly encountered in milk samples.

Elevated cardiac troponin levels predict the risk of adverse outcome in patients with acute coronary syndromes.

BACKGROUND: Elevations of cardiac troponin T or I are predictive of adverse outcomes in patients with acute coronary syndromes. However, odds ratios (ORs) vary substantially between studies. This investigation refines these values by means of a meta-analysis. METHODS: Twenty-one studies were suitable. ORs were calculated for short-term (30 days) and long-term (5 months to 3 years) follow-up in patients with ST-segment elevation (ST upward arrow), in those without ST-segment elevation (no ST upward arrow), and in patients with unstable angina. The primary end point was a composite of death or nonfatal myocardial infarction. RESULTS: A total of 18,982 patients were included. At 30 days, the OR for death or myocardial infarction was 3.44 (95% confidence interval [CI], 2.94-4.03; P <. 00001) for patients with positive troponin. In the ST upward arrow group, troponin elevations carried a 2.86-fold (95% CI, 2.35-3.47; P <.0001) higher risk during short-term follow-up, which was maintained long term. The no-ST upward arrow patients with troponin elevations manifested a 4.93-fold (95% CI, 3.77-6.45; P <.0001) increase of adverse outcomes. The OR for patients with unstable angina and positive troponin was 9.39 (95% CI, 6.46-13.67; P <.0001). For cardiac death alone, the results were similar. CONCLUSIONS: Patients with acute coronary syndromes who have troponin elevations show a substantial increase in risk during short and long-term follow-up.

Two-dimensional gel electrophoresis/matrix-assisted laser desorption/ionisation mass spectrometry of a milk powder.

Proteins in a commercial milk powder have been separated by two-dimensional gel electrophoresis and analysed by matrix-assisted laser desorption ionisation mass spectrometry. The mass spectrometric analyses were conducted in two steps: analysis of the intact proteins following their passive extraction into a suitable solvent mixture and analysis in reflectron mode of in situ digests of a number of gel spots. The combination of the two methods allowed a reliable identification of a number of proteins, including nine caseins as well as certain protein modifications including single/multiple phosphorylation, lactose-protein conjugates and Coomassie Brilliant Blue adducts. Analyses of the intact proteins prior to their in situ digestion contributed to a more efficient and reliable consultation of protein databases.

Probing protein unfolding through monitoring cysteine alkylation by matrix-assisted laser desorption/ionisation mass spectrometry.

Matrix-assisted laser desorption/ionisation mass spectrometry was used to monitor interaction between three proteins and two basic Immobiline chemicals (pK 10.3 and pK >12) commonly used in immobilised pH gradients (IPG). For two of the investigated proteins, the observed alkylation channels of the cysteine residues exhibited unmistakable response to their gradual denaturation following treatment with different concentrations (0-8 M) of two commonly used denaturants, urea and guanidine hydrochloride. Our assessment for protein unfolding is based on the number and relative intensity of the alkylation channels, yet the present mass spectrometry data are in good agreement with data based on optical rotatory dispersion, in which another approach was used to assess protein unfolding. Whether the present simple, fast and specific mass spectrometry method can be developed as a probe for monitoring folding/unfolding of cysteine-containing proteins can only be demonstrated by generating similar data for a larger number of proteins.