Esteban Orenes-Piñero

Small-size circulating microparticles in acute coronary syndromes: relevance to fibrinolytic status, reparative markers and outcomes.

BACKGROUND Recent evidence suggests that circulating microparticles (MPs) contribute to inflammation, coagulation and vascular injury. Majority of MPs have usually not been included into prior analyses due their small size and limited resolution of conventional equipment. Our aim was to assess levels of MPs of different cellular origin sized below 0.5 μm polystyrene beads, denoted as small-size microparticles (sMP), their relation to markers of cardiovascular repair and their impact on prognosis in patients with acute coronary syndromes (ACS). METHODS In a cross-sectional study, we initially compared levels of sMP between patients with ST-segment elevation myocardial infarction (STEMI, n = 50), non-STEMI (n = 47), stable coronary artery disease (CAD, n = 40) and healthy individuals (HC, n = 40). In a separate study, the prognostic value of sMP was assessed in patients with non-STEMI (n = 160). Annexin V-binding sMP (sAMP), platelet CD42b(+) sMPs (sPMP), endothelial CD144(+) sMP (sEMP) and monocyte CD14(+) sMP (sMMP) were quantified using high resolution flow cytometry. Endothelial progenitor cells (EPCs) and monocyte expression of scavenger receptors was quantified by flow cytometry. Fibrinolytic factors were measured by ELISA. RESULTS Counts of sAMP and sEMP were lower in STEMI after PCI (p < 0.001 and p = 0.025, respectively) but not in non-STEMI vs. CAD. sAMP was positively correlated with EPCs in non-STEMI (p < 0.001). Likewise, plasminogen activators negatively correlated with sAMP in non-STEMI and STEMI (p = 0.02 and p = 0.002, respectively). In non-STEMI patients, sEMP and sMMP were independently predictive for future admissions related to heart failure (p = 0.034 and 0.013, respectively) and sPMP for major bleedings (p = 0.002). The sAMP/EPCs ratio was higher in patients (before PCI) compared to STEMI patients. CONCLUSIONS Small-size MPs could be potentially implicated in the modulation of the post-ACS reparative response to injury, with prognostic implications. Besides, the sAMP/EPCs ratio could reflect a change in the apoptotic/reparative potential, being a putative indicator for vascular repair.

Controversies Surrounding High-Protein Diet Intake: Satiating Effect and Kidney and Bone Health

Long-term consumption of a high-protein diet could be linked with metabolic and clinical problems, such as loss of bone mass and renal dysfunction. However, although it is well accepted that a high-protein diet may be detrimental to individuals with existing kidney dysfunction, there is little evidence that high protein intake is dangerous for healthy individuals. High-protein meals and foods are thought to have a greater satiating effect than high-carbohydrate or high-fat meals. The effect of high-protein diets on the modulation of satiety involves multiple metabolic pathways. Protein intake induces complex signals, with peptide hormones being released from the gastrointestinal tract and blood amino acids and derived metabolites being released in the blood. Protein intake also stimulates metabolic hormones that communicate information about energy status to the brain. Long-term ingestion of high amounts of protein seems to decrease food intake, body weight, and body adiposity in many well-documented studies. The aim of this article is to provide an extensive overview of the efficacy of high protein consumption in weight loss and maintenance, as well as the potential consequences in human health of long-term intake.

An innovative flow cytometric approach for small-size platelet microparticles: Influence of calcium

Microparticles (MPs) are small submicron membrane-derived vesicles shed from a variety of cells and they have been implicated in different disorders. Accordingly, understanding of physiological characteristics of MPs and improvement of methods of their quantification are important for further advance in the field. Although flow cytometry is the most widely applied technique for MP analysis, it is limited by lack of adequate standardisation. Annexin V (AnV), which binds surface phosphatidylserine (PS) with high affinity, has been long regarded as a marker of MPs, but AnV binding is Ca2+-dependent and it is unclear how [Ca2+] concentrations could affect AnV binding to MPs and its enumeration. MPs from citrated and heparinised plasma were labelled with AnV, anti-CD42b and quantified using an Apogee A50 flow cytometer. The small-size MP gate was defined with the use of size beads (from 0.1 to 0.5 μm) and confirmed with an in vitro assessment of platelet stimulation. Biotinylated anti-CD42b antibodies were then bound to streptavidin conjugated with different fluorochromes, leading to an amplified signal of platelet MPs (PMPs). Moderate increase of [Ca2+] concentrations in the annexin V staining buffer allows initial plasma recalcification and more accurate MP quantification in citrated plasma. Thrombin stimulation of platelet-free plasma containing only MPs did not produce any changes in the concentration of AnV+ MPs, but decreased the anti-CD42b binding. The results also indicate that prolonged storage and thrombin induce the release of AnV+ MPs whereas PS exposure in pre-existent MPs is not affected by thrombin. In conclusion, we present a sensitive protocol for the analysis of circulating and in vitro induced small-size PMPs that might contribute to future cardiovascular and clinical research.

β-Trace Protein: From GFR Marker to Cardiovascular Risk Predictor

β-Trace protein, also known as Lipocalin type prostaglandin D synthase, is a low-molecular mass glycoprotein (between 23,000 and 29,000 Da depending on the degree of glycosylation) that converts prostaglandin H2 into prostaglandin D2. β-Trace protein was initially isolated from cerebrospinal fluid and served as a marker of cerebrospinal fluid leakage; however, its cDNA and gene have been isolated in numerous human body tissues, including central nervous system, retina, melanocytes, heart, and male genital organs. In recent years, β-trace protein has emerged as a promising novel endogenous marker of GFR, representing a more sensitive marker for mild kidney dysfunction than serum creatinine. In this regard, β-trace protein has been proposed as an alternative marker to Cystatin C for measuring kidney function. Beyond its role for estimating renal function, β-trace protein is also emerging as a novel biomarker in cardiovascular risk. It has been associated with several cardiovascular disorders, playing a potential role for prognostic stratification in patients with acutely decompensated heart failure and acute coronary syndromes and being advocated as a novel marker for cardiovascular risk prediction.

Anti-inflammatory Effects of Omega 3 and Omega 6 Polyunsaturated Fatty Acids in Cardiovascular Disease and Metabolic Syndrome.

ABSTRACT A lipid excess produces a systemic inflammation process due to tumor necrosis factor-α, interleukin-6 and C-reactive protein synthesis. Simultaneously, this fat excess promotes the appearance of insulin resistance. All this contributes to the development of atherosclerosis and increases the risk of cardiovascular diseases (CVDs). On the other hand, polyunsaturated fatty acids (PUFAs), especially eicosapentaenoic acid and docosahexaenoic acid (omega 3), and arachidonic acid (omega 6) have shown anti-inflammatory properties. Lately, an inverse relationship between omega-3 fatty acids, inflammation, obesity and CVDs has been demonstrated. To check fatty acids effect, the levels of some inflammation biomarkers have been analyzed. Leptin, adiponectin and resistin represent a group of hormones associated with the development of CVDs, obesity, type 2 diabetes mellitus and insulin resistance and are modified in obese/overweight people comparing to normal weight people. Omega-3 PUFAs have been shown to decrease the production of inflammatory mediators, having a positive effect in obesity and diabetes mellitus type-2. Moreover, they significantly decrease the appearance of CVD risk factors. Regarding omega-6 PUFA, there is controversy whether their effects are pro- or anti-inflammatory. The aim of this manuscript is to provide a comprehensive overview about the role of omega-3 and omega-6 PUFAs in CVDs and metabolic syndrome.

Comparison of estimated glomerular filtration rate equations for dosing new oral anticoagulants in patients with atrial fibrillation

INTRODUCTION AND OBJECTIVES New oral anticoagulants require dosing adjustment according to renal function. We aimed to determine discordance in hypothetical recommended dosing of these drugs using different estimated glomerular filtration rate equations in patients with atrial fibrillation. METHODS Cross-sectional analysis of 910 patients with atrial fibrillation and an indication for oral anticoagulation. The glomerular filtration rate was estimated using the Cockcroft-Gault, Modification of Diet in Renal Disease and Chronic Kidney Disease Epidemiology Collaboration equations. For dabigatran, rivaroxaban, and apixaban we identified dose discordance when there was disagreement in the recommended dose based on different equations. RESULTS Among the overall population, relative to Cockcroft-Gault, discordance in dabigatran dosage was 11.4% for Modification of Diet in Renal Disease and 10% for Chronic Kidney Disease Epidemiology Collaboration, discordance in rivaroxaban dosage was 10% for Modification of Diet in Renal Disease and 8.5% for the Chronic Kidney Disease Epidemiology Collaboration. The lowest discordance was observed for apixaban: 1.4% for Modification of Diet in Renal Disease and 1.5% for the Chronic Kidney Disease Epidemiology Collaboration. In patients with Cockcroft-Gault<60mL/min or elderly patients, discordances in dabigatran and rivaroxaban dosages were higher, ranging from 13.2% to 30.4%. Discordance in apixaban dosage remained<5% in these patients. CONCLUSIONS Discordance in new oral anticoagulation dosages using different equations is frequent, especially among elderly patients with renal impairment. This discordance was higher in dabigatran and rivaroxaban dosages than in apixaban dosages. Further studies are needed to clarify the clinical importance of these discordances and the optimal anticoagulant dosages depending on the use of different equations to estimate renal function.

High-sensitivity troponin T as a biomarker for the development of atrial fibrillation after cardiac surgery.

OBJECTIVES Atrial fibrillation (AF) occurs in ∼ 30% of patients undergoing coronary artery bypass grafting (CABG) and in 40% of patients after valve surgery. High-sensitivity cardiac troponin T (hsTnT) is a specific and high-sensitivity marker of myocardial injury, while N-terminal proB-type natriuretic peptide (NT-proBNP) is an established biomarker for wall remodelling. We investigated whether hsTnT and NT-proBNP levels could be used as valuable biomarkers for AF occurrence after cardiac surgery. METHODS We included consecutive haemodynamically stable patients undergoing programmed cardiac surgery with cardiopulmonary bypass pump. We determined hsTnT and NT-proBNP levels before and after cardiac surgery and recorded AF development by prolonged electrocardiogram monitoring. RESULTS We included 100 patients with predominantly aortic valve (n = 42) or ischaemic heart (n = 58) diseases. Twenty-nine patients (29%) developed post-surgical AF. Patients developing AF had a longer hospital stay (P = 0.005). hsTnT levels increased after surgery [P < 0.001], indicating perioperative myocardial injury, with higher presurgery levels in patients who developed AF [P = 0.015]. Body mass index and EuroSCORE risk scale were independently associated with higher hsTnT levels presurgery. On univariate analysis, age (P = 0.048), male sex (P = 0.031), indexed left atrial volume (P = 0.042), β-blockers treatment (P = 0.024), type of surgery (valve surgery vs CABG; P = 0.034), EuroSCORE risk scale (P = 0.025) and higher preoperative hsTnT levels (P = 0.009) were predictors of AF development, but NT-proBNP did not reach statistical significance (P = 0.060). hsTnT levels in blood samples obtained the day after surgery were not associated with post-surgical AF development (P = 0.165). In a multivariate model, only higher hsTnT levels before cardiac surgery (>11.87 ng/l) [Odds Ratio, OR; (95% Confidence interval, CI) 4.27 (1.43-12.77), P = 0.009] and male sex [OR 5.10 (1.72-15.13), P = 0.003)] were independently associated with the occurrence of post-surgical AF. CONCLUSION High presurgical hsTnT levels were independently predictive of patients developing AF after cardiac surgery. hsTnT levels determined post-surgery suggest that cardiac perioperative myocardial injury is not associated with postoperative AF development. NT-proBNP did not reach statistical significance as a biomarker for AF prediction.

Familial nonmedullary thyroid cancer: Screening, clinical, molecular and genetic findings

Thyroid cancer, the commonest of endocrine malignancies, continues increasing in incidence being the 5th more prevalent cancer among women in the United States in 2012. Familial thyroid cancer has become a well-recognized, unique, clinical entity in patients with thyroid cancer originating from follicular cells, that is, nonmedullary thyroid carcinoma. Hereditary nonmedullary thyroid cancer may occur as a minor component of familial cancer syndromes (familial adenomatous polyposis, Gardners syndrome, Cowdens disease, Carneys complex type 1, Werners syndrome, and papillary renal neoplasia) or as a primary feature (familial nonmedullary thyroid cancer [FNMTC]). Although there is some controversy, some epidemiologic and clinical kindred studies have shown that FNMTC is associated with more aggressive disease than sporadic cases, with higher rates of multicentric tumours, lymph node metastasis, extrathyroidal invasion, and shorter disease-free survival. This way, preventing screening will allow earlier detection, more timely intervention, and hopefully improved outcomes for patients and their families. On the other hand, in the last years, an important number of genetic studies on FNMTC have been published, trying to determine its genetic contribution. However, the genetic inheritance of FNMTC remains unclear; but it is believed to be autosomal dominant with incomplete penetrance and variable expressivity. This paper provides an extensive overview of FNMTC from several points of view. Firstly, the impact of early detection on prognosis, secondly, the management and follow-up of FNMTC patients, and finally, the role of susceptibility loci, microRNAs (miRNAs) and telomerases in recently identified isolated cases of FNMTC.

A new process for obtaining hydroxytyrosol using transformed Escherichia coli whole cells with phenol hydroxylase gene from Geobacillus thermoglucosidasius

Phenol hydroxylase gene cloning from the thermophilic bacteria Geobacillus thermoglucosidasius was used to develop an effective method to convert tyrosol into the high-added-value compound hydroxytyrosol by hydroxylation. Phenol hydroxylase is a two-component enzyme encoded by pheA1 and pheA2 genes and strictly dependent on NADH and FAD. These two genes were subcloned together as a 2 kb fragment into Escherichia coli Rosetta cells, and the transformants were able to grow and effectively transform up to 5 mM of phenol and tyrosol using IPTG (isopropyl-β-D-thiogalactopyranoside) as inducer. In addition, when a new fragment with a 340 pb upstream pheA1 gene was subcloned, a similar biotransformation rate was attained without IPTG, confirming that this fragment encodes for a phenol hydroxylase promoter that can be recognised by E. coli. Both transformants brought about the total bioconversion of monophenols at a high concentration (5 mM), which represents an increase, both in concentration and in yield, compared with that previously described in the bibliography. The use of the transformant with its constitutive promoter was more interesting from a biotechnological point of view, since it is not necessary to use IPTG. It also gave rise to greater operational stability.

Usefulness of β-trace protein and cystatin C for the prediction of mortality in non ST segment elevation acute coronary syndromes.

Beta-trace protein (BTP) is a low-molecular mass protein belonging to the lipocalin protein family, which is more sensitive than serum creatinine for detecting impaired renal function. The aims of the present study were to evaluate whether plasma BTP improves the risk stratification of patients with non-ST-segment elevation acute coronary syndromes and to compare it to cystatin C (CysC), serum creatinine, and estimated glomerular filtration rate. Two hundred twenty-six consecutive patients with non-ST-segment elevation acute coronary syndromes were prospectively included. Blood samples were obtained within 24 hours of hospital admission to measure BTP, CysC, and creatinine. The study end point was all-cause death. Over a median follow-up period of 859 days (interquartile range [IQR] 524 to 1,164), 24 patients (10.6%) died. Decedents had higher concentrations of BTP (1.03 mg/L [IQR 0.89 to 1.43] vs 0.74 mg/L [IQR 0.61 to 0.92], p <0.001), CysC (1.16 mg/L [IQR 0.91 to 1.59] vs 0.90 mg/L [IQR 0.76 to 1.08], p = 0.001), and serum creatinine (1.10 mg/L [IQR 0.87 to 1.46] vs 0.94 mg/L [IQR 0.80 to 1.10], p = 0.004) and a lower mean estimated glomerular filtration rate (60 ± 20 vs 80 ± 24 ml/min/1.73 m(2), p <0.001). After multivariate adjustment, BTP and CysC were predictors of all-cause death, while estimated glomerular filtration rate and serum creatinine concentrations did not achieve statistical significance. In stratified analyses according to kidney function, elevated BTP and CysC were associated with a higher risk for all-cause death. Reclassification analyses showed that BTP and CysC added complementary information to Global Registry for Acute Coronary Events (GRACE) risk score. In conclusion, BTP and CysC levels were associated with all-cause death risk and modestly improved prognostic discrimination beyond the GRACE risk score in patients with non-ST segment elevation acute coronary syndromes.