Glycation of human serum albumin impairs binding to the glucagon-like peptide-1 analogue liraglutide.

The long-acting glucagon-like peptide-1 analogue liraglutide has proven efficiency in the management of type 2 diabetes and also has beneficial effects on cardiovascular diseases. Liraglutide's protracted action highly depends on its capacity to bind to albumin via its palmitic acid part. However, in diabetes, albumin can undergo glycation, resulting in impaired drug binding. Our objective in this study was to assess the impact of human serum albumin (HSA) glycation on liraglutide affinity. Using fluorine labeling of the drug and 19F NMR, we determined HSA affinity for liraglutide in two glycated albumin models. We either glycated HSA in vitro by incubation with glucose (G25- or G100-HSA) or methylglyoxal (MGO-HSA) or purified in vivo glycated HSA from the plasma of diabetic patients with poor glycemic control. Nonglycated commercial HSA (G0-HSA) and HSA purified from plasma of healthy individuals served as controls. We found that glycation decreases affinity for liraglutide by 7-fold for G100-HSA and by 5-fold for MGO-HSA compared with G0-HSA. A similarly reduced affinity was observed for HSA purified from diabetic individuals compared with HSA from healthy individuals. Our results reveal that glycation significantly impairs HSA affinity to liraglutide and confirm that glycation contributes to liraglutide's variable therapeutic efficiency, depending on diabetes stage. Because diabetes is a progressive disease, the effect of glycated albumin on liraglutide affinity found here is important to consider when diabetes is managed with this drug.

The different roles for the advanced glycation end products axis in heart failure and acute coronary syndrome settings.

AIMS: This work aimed to compare the behavior of the advanced glycation end products (AGEs) and their soluble receptor (sRAGE) in two cohorts of patients: those with heart failure (HF) and acute coronary syndrome (ACS). METHODS AND RESULTS: A unicentric observational clinical study was performed in 102 patients with ACS and 102 patients with chronic HF matched by age and gender. At inclusion, fluorescent AGEs were measured by quantitative fluorescence spectroscopy of plasma, and total sRAGE and endogenous secretory RAGE (esRAGE) levels were determined by enzyme-linked immunosorbent assay kits. A 5-year follow-up period was established for recording cardiac death (primary endpoint) and the incidence of non-fatal myocardial infarction or HF readmission (secondary endpoints). Higher glycation parameters were observed in HF patients, whereas no differences in sRAGE forms were found between HF and ACS cohorts, except for cRAGE, which was higher in HF. Associations between glycation parameters and sRAGE forms were observed in HF, but not in ACS. Differences were also evidenced in the long-term prognosis of each cohort: esRAGE showed an independent prognostic value for cardiac death or non-fatal cardiovascular events in HF, but none of the AGE-RAGE variables were predictors in ACS. CONCLUSIONS: A different role for the AGE-RAGE axis was observed in HF and ACS. All the sRAGE forms were directly related with glycation parameters in HF, but not in ACS. The independent value of the sRAGE forms on each cardiovascular disease was supported by esRAGE being an independent predictor of bad long-term prognosis only for HF.

Protective, repairing and fibrinolytic effects of rivaroxaban on vascular endothelium.

AIMS: Rivaroxaban, a direct inhibitor of activated factor X (FXa), is the only new oral anticoagulant approved for secondary prevention after acute coronary syndrome. Our objective was to identify the possible molecular mechanisms of rivaroxaban that contribute to endothelial function. METHODS: Cell viability and growth of human umbilical vein endothelial cells (HUVEC) were registered. Gene expression studies comparing the effects of rivaroxaban and FXa were conducted by a selective RNA array and confirmed by protein quantification. Wound-healing experiments on HUVEC, platelet adhesion, enzymatic activity, and cell-based assays for fibrin formation were performed with rivaroxaban. RESULTS: Rivaroxaban (50 nM) only altered (>2 fold change) the expression of matrix metallopeptidase 2 and urokinase plasminogen activator (u-PA), but counteracted the FXa (9 nM)-induced upregulation of several pro-inflammatory genes (P < 0.05) and FXa-enhanced platelet adhesion over HUVEC. Rivaroxaban increased u-PA protein expression in HUVEC supernatants and enhanced u-PA activity (up to 4 IU ng-1 of u-PA). Rivaroxaban (1 nM-1 µM) showed a significant and dose-dependent positive effect on HUVEC growth that was inhibited by BC-11-hydroxibromide, an inhibitor of u-PA. Healing properties after a wound on HUVEC cultures, and fibrinolytic properties were also shown by rivaroxaban. Both effects were reversed by BC-11-hydroxibromide. CONCLUSIONS: Rivaroxaban enhanced viability, growth and migration of HUVEC, mainly by u-PA activation and upregulation, which also participate in the rivaroxaban-induced fibrinolytic activity at endothelial level. Rivaroxaban also protected from the pro-inflammatory effects of FXa on HUVEC. Altogether may improve endothelial functionality and could contribute to the cardiovascular benefits of rivaroxaban.

Evolution and bad prognostic value of advanced glycation end products after acute heart failure: relation with body composition.

AIM: The role of advanced glycation end products (AGEs) and their soluble receptor (sRAGE) on the progression and prognosis of acute heart failure (HF) was analysed in relation with metabolic parameters as body composition and nutritional status. METHODS: A hundred and fifty consecutive patients were included in a prospective clinical study during hospitalization by acute HF. Detailed medical history, physical examination, electrocardiogram, echocardiogram and vein peripheral blood were taken for all patients. During the follow-up period [297 days (88-422 days)] blood samples for biochemical measurements were obtained 1 and 6 months after the inclusion. Dual-energy X-ray absorptiometry analyses were performed 1 week after discharge. RESULTS: AGEs and sRAGE levels continuously increased, up to 6 months, after acute HF, but AGEs increase was mainly observed in those patients with incident HF. Both AGEs and sRAGE levels were related with bad renal function and clinical malnutrition (CONUT score) and they were negatively related with body mass index or percentage of body fat. AGEs levels (≥40 a.u.) 1 month after discharge and basal sRAGE levels (>1000 pg/mL) were related with worse prognosis in terms of patient death and HF readmission (Log-rank <0.05 in Kaplan-Meier survival test), independently of age, gender, body mass index and other risk factors. Regression models also corroborated this finding. CONCLUSIONS: AGEs and sRAGE are bad prognostic biomarkers for HF and useful markers of HF progression. Since their levels seem to be related with clinical malnutrition and body composition these parameters could serve to modulate them.

Glucose and Inflammatory Cells Decrease Adiponectin in Epicardial Adipose Tissue Cells: Paracrine Consequences on Vascular Endothelium.

Epicardial adipose tissue (EAT) is a source of energy for heart that expresses the insulin-sensitizer, anti-inflammatory and anti-atherogenic protein, adiponectin. But, in coronary artery disease, adiponectin production declines. Our objective was to determine its regulation by glucose and inflammation in stromal cells from EAT and subcutaneous adipose tissue (SAT) and its paracrine effect on endothelial cells. Stromal cells of EAT and SAT were obtained from patients who underwent cardiac surgery. Adipogenesis was induced at 117, 200, or 295 mg/dl glucose, with or without macrophage-conditioned medium (MCM). Expression of adiponectin, GLUT-4 and the insulin receptor was analyzed by real-time PCR. The paracrine effect of stromal cells was determined in co-cultures with endothelial cells, by exposing them to high glucose and/or MCM, and, additionally, to leukocyte-conditioned medium from patients with myocardial infarction. The endothelial response was determined by analyzing vascular adhesion molecule expression. Our results showed a U-shaped dose-response curve of glucose on adiponectin in EAT, but not in SAT stromal cells. Conversely, MCM reduced the adipogenesis-induced adiponectin expression of EAT stromal cells. The presence of EAT stromal increased the inflammatory molecules of endothelial cells. This deleterious effect was emphasized in the presence of inflammatory cell-conditioned medium from patients with myocardial infarction. Thus, high glucose and inflammatory cells reduced adipogenesis-induced adiponectin expression of EAT stromal cells, which induced an inflammatory paracrine process in endothelial cells. This inflammatory effect was lower in presence of mature adipocytes, producers of adiponectin. These results contribute to understanding the role of EAT dysfunction on coronary atherosclerosis progression.

Endothelial progenitor cells mobilisation after percutaneous coronary intervention: a pilot study.

BACKGROUND: The mobilisation process of endothelial progenitor cells (EPC) after stent implantation by percutaneous coronary intervention (PCI) is unclear because the circulating EPC levels are influenced by several pathophysiological factors. The objective was to analyse the kinetics of EPC concentration following elective PCI in patients with stable angina, and its relation with other biomarkers or parameters of cardiovascular function. METHODS: Pilot study in stable angina patients (n = 30) for elective PCI and implantation of bare-metal stent (BMS), drug-eluting stent (DES) or EPC-capturing stent (ECS). Samples were taken at baseline, 6 h, 24 h and 6 months after PCI for biochemical analysis and EPC quantification by flow cytometry. RESULTS: Baseline EPC levels, quantified in peripheral blood, were related with the extent of the coronary lesion and the percentage of stenosis. EPC concentration increased 6 hours after PCI in relation with plasma C-reactive protein concentration and returned to basal levels after 24 hours post-PCI. CONCLUSIONS: Baseline EPC levels are related with the extension of the lesion and stenosis whereas the kinetics of EPC mobilization showed to be related with C-reactive protein concentration. Endothelial activation seems to occur in response to EPC mobilization or vascular damage by PCI.

High-sensitivity C-reactive protein predicts adverse outcomes after non-ST-segment elevation acute coronary syndrome regardless of GRACE risk score, but not after ST-segment elevation myocardial infarction.

INTRODUCTION: Atherosclerosis is an active process and the inflammatory component appears to be particularly correlated with the development of acute coronary syndromes (ACS). C-reactive protein (CRP) is an acute phase protein that appears in the circulation in response to inflammatory cytokines. The present study investigated the association between high-sensitivity C-reactive protein (hsCRP) on admission and follow-up prognosis after an ACS. METHODS: We included 151 consecutive patients admitted to the coronary care unit with a diagnosis of ACS (47% ST-segment elevation myocardial infarction [STEMI]). The primary endpoint was the combination of cardiac death and myocardial reinfarction during the follow-up period (median 19.8 months, interquartile range 16.3-23.7 months). RESULTS: The occurrence of follow-up events was significantly related to admission hsCRP level, which was an excellent predictor of cardiac death and reinfarction during follow-up (HR 1.091, 95% CI 1.014-1.174; p=0.019). Stratifying the population based on type of ACS, adjusted by variables associated with cardiac events in univariate analysis (hsCRP, diabetes, depressed ejection fraction and GRACE risk score), hsCRP proved to be an independent predictor of follow-up outcomes only in non-STEMI patients (HR 1.217, 95% CI: 1.093-1.356, p<0.001), not in STEMI patients. The best cutoff level of hsCRP to predict follow-up outcomes was 1.1mg/dl, with sensitivity of 77.8% and specificity of 63.2%. CONCLUSION: Although the GRACE risk score is routinely used for stratification of patients with ACS, assessment of hsCRP may provide additional prognostic value in the follow-up of non-STEMI patients.

Predictive value of advanced glycation end products for the development of post-infarction heart failure: a preliminary report.

BACKGROUND: Since post-infarction heart failure (HF) determines a great morbidity and mortality, and given the physiopathology implications of advanced glycation end products (AGE) in the genesis of myocardial dysfunction, it was intended to analyze the prognostic value of these molecules in order to predict post-infarction HF development. METHODS: A prospective clinical study in patients after first acute coronary syndrome was conducted. The follow-up period was consisted in 1 year. In 194 patients consecutively admitted in the coronary unit for myocardial infarct fluorescent AGE levels were measured. The association between glycaemic parameters and the development of post-infarction HF were analyzed in those patients. Finally, we identified the variables with independent predictor value by performing a multivariate analysis of hazard ratio for Cox regression. RESULTS: Eleven out of 194 patients (5.6%) developed HF during follow-up (median: 1.0 years [0.8 - 1.5 years]). Even though basal glucose, fructosamine and glycated haemoglobin were significant predictive factors in the univariate analysis, after being adjusted by confounding variables and AGE they lost their statistical signification. Only AGE (Hazard Ratio 1.016, IC 95%: 1.006-1.026; p<0,001), together with NT-proBNP and the infarct extension were predictors for post-infarction HF development, where AGE levels over the median value 5-fold increased the risk of HF development during follow-up. CONCLUSIONS: AGE are an independent marker of post-infarction HF development risk.

Galectin-3 and soluble RAGE as new biomarkers of post-infarction cardiac remodeling.

Post-infarction remodeling is a clinical problem with no curative treatment. Our objective was to search for new biomarkers of cardiac remodeling that have clinical value after ST-segment elevation myocardial infarction (STEMI). This pilot study enrolled 67 consecutive patients with de novo STEMI who underwent revascularization by primary angioplasty. Echocardiography studies of cardiac function were completed during the first 48 h post-STEMI and after 6 months of follow-up. Galectin-3 and soluble receptor for advanced glycation end products (sRAGE) were tested in the peripheral venous blood during the 24 h post-infarction. Cardiac remodeling was defined as changes ≥ 15% in the left ventricular end-systolic volume (LVESV) or > 10% in the left atrial area (LAA). An inverse association was found between galectin-3 (rs = - 0.296; p < 0.001) and sRAGE (rs = - 0.327; p < 0.001) levels and the basal left ventricle ejection fraction (LVEF). However, only galectin-3 was directly associated with the increase in LVESV (rs = 0.389; p = 0.007) and LVEDV (rs = 0.314; p = 0.031) during the follow-up. sRAGE was inversely related to the change in LAA (rs = - 0.320; p = 0.032). These data are consistent with galectin-3, but not sRAGE levels, as a predictor of left ventricle remodeling (OR 1.036, 95% CI 1.002-1.071; p = 0.039). Galectin-3 and sRAGE levels that were measured during hospitalization are inversely related to basal LVEF after a STEMI. Galectin-3 levels are a predictor of adverse post-STEMI LV remodeling, whereas sRAGE levels exhibited an inverse relationship with left atrial remodeling. KEY MESSAGES: Post-infarction remodeling is a clinical problem with no curative treatment. New biomarkers for remodeling after acute myocardial infarction were explored. Early post-STEMI galectin-3 and soluble RAGE are inversely related with left ventricle function. Galectin-3 levels were predictors of adverse post-STEMI left ventricle remodeling. Soluble RAGE levels were associated with left atrial remodeling.

Inflammatory effects of in vivo glycated albumin from cardiovascular patients.

OBJECTIVES: Characterization of the type of glycation found in circulating proteins from cardiovascular patients in comparison with healthy control subjects and to explore the pathophysiological molecular effects of these glycomodified proteins on human umbilical vein endothelial cells (HUVEC) in culture. METHODS: Human serum albumin pools from 10 subjects each, of patients with heart failure (HF) presenting high or low glycation levels, and from healthy subjects were isolated and purified. The glycation levels of these pools were characterized by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and compared between them. Analysis of endothelial dysfunction after the treatment of HUVEC with the pools was made by mRNA expression of adhesion molecules and by functional adhesion of mononuclear cells to HUVEC monolayers. RESULTS: Specific characterization of post-transductional modifications (advanced glycation end products) in high and low glycated albumins from patients was made in comparison with healthy subjects. Albumins from patients were able, at very low concentrations (12.5 µg/mL), to significantly up-regulate (˜0.2 - 2 fold) the gene expression of adhesion molecules in HUVEC. At the functional level, the albumin from patients with high glycation levels (at 12.5 and 25 µg/mL) significantly enhanced (˜10%) the adhesion of mononuclear cells to HUVEC. CONCLUSIONS: Differences in the glycomodification of albumin from HF patients were found and specifically characterized in comparison with albumin from healthy subjects. Functionally, in vivo glycated albumin in patients with HF induced an increase in adhesion molecules expression on HUVEC, which supported an increase in peripheral blood mononuclear cells adhesion to endothelial cells.

Advanced glycation end-products disrupt human endothelial cells redox homeostasis: new insights into reactive oxygen species production.

Advanced glycation end-products (AGEs) trigger multiple metabolic disorders in the vessel wall that may in turn lead to endothelial dysfunction. The molecular mechanisms by which AGEs generate these effects are not completely understood. Oxidative stress plays a key role in the development of deleterious effects that occur in endothelium during diabetes. Our main objectives were to further understand how AGEs contribute to reactive oxygen species (ROS) overproduction in endothelial cells and to evaluate the protective effect of an antioxidant plant extract. The human endothelial cell line EA.hy926 was treated with native or modified bovine serum albumin (respectively BSA and BSA-AGEs). To monitor free radicals formation, we used H2DCF-DA, dihydroethidium (DHE), DAF-FM-DA and MitoSOX Red dyes. To investigate potential sources of ROS, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and mitochondrial inhibitors were used. The regulation of different types of ROS by the polyphenol-rich extract from the medicinal plant Doratoxylon apetalum was also studied for a therapeutic perspective. BSA-AGEs exhibited not only less antioxidant properties than BSA, but also pro-oxidant effects. The degree of albumin glycoxidation directly influenced oxidative stress through a possible communication between NADPH oxidase and mitochondria. D. apetalum significantly decreased intracellular hydrogen peroxide and superoxide anions mainly detected by H2DCF-DA and DHE respectively. Our results suggest that BSA-AGEs promote a marked oxidative stress mediated at least by NADPH oxidase and mitochondria. D. apetalum plant extract appeared to be an effective antioxidant compound to protect endothelial cells.

Diabetes-induced hepatic oxidative stress: a new pathogenic role for glycated albumin.

Increased oxidative stress and advanced glycation end-product (AGE) formation are major contributors to the development of type 2 diabetes. Here plasma proteins e.g. albumin can undergo glycoxidation and play a key role in diabetes onset and related pathologies. However, despite recent progress linking albumin-AGE to increased oxidative stress and downstream effects, its action in metabolic organs such as the liver remains to be elucidated. The current study therefore investigated links between oxidative perturbations and biochemical/structural modifications of plasma albumin, and subsequent downstream effects in transgenic db/db mouse livers and HepG2 cells, respectively. Our data reveal increased oxidative stress biomarkers and lipid accumulation in plasma and livers of diabetic mice, together with albumin glycoxidation. Purified mouse albumin modifications resembled those typically found in diabetic patients, i.e. degree of glycation, carbonylation, AGE levels and in terms of chemical composition. Receptor for AGE expression and reactive oxygen species production were upregulated in db/db mouse livers, together with impaired proteolytic, antioxidant and mitochondrial respiratory activities. In parallel, acute exposure of HepG2 cells to glycated albumin also elicited intracellular free radical formation. Together this study demonstrates that AGE-modified albumin can trigger damaging effects on the liver, i.e. by increasing oxidative stress, attenuating antioxidant capacity, and by impairment of hepatic proteolytic and respiratory chain enzyme activities.

Sea cucumbers with an anti-inflammatory effect on endothelial cells and subcutaneous but not on epicardial adipose tissue.

OBJECTIVE: epicardial adipose tissue (EAT) from patients with coronary artery disease (CAD) contains higher levels of inflammatory proteins and lower adiponectin levels than subcutaneous adipose tissue (SAT), enhancing the progression of atherosclerosis. Since products from sea cucumber have anti-inflammatory properties, we investigated its effect on EAT, SAT and endothelial cells. METHODS: stromal cells or explants from EAT and SAT were obtained from patients with cardiovascular disease. Extracts were obtained after hydrolysis by food-grade enzymes at different times. Proteins were identified by LC-MALDI mass spectrometry. Adipogenesis and adiponectin induction were determined on stromal cells in the presence/absence of extracts. The bioavailability of the extracts was tested on a Caco-2 cell culture model in vitro. The bioavailable fraction was probed on endothelial cells and EAT or SAT explants. Vascular cell adhesion protein (VCAM-1), intercellular adhesion molecule (ICAM-1), IL-6 and adiponectin were determined by real time polymerase chain reaction (RT-PCR). RESULTS: our results showed that H. forskali and P. tremulus extracts contained compounds with anti-oxidant and anti-inflammatory properties. The bioavailable fraction of P. tremulus reduced VCAM-1 (p < 0.01) and IL-6 (p < 0.05) expression levels in endothelial cells while bioavailable compounds from H. forskali decreased ICAM-1 expression in SAT (p < 0.05). No effect was observed on EAT. CONCLUSION: these results suggest that sea cucumber extracts might be used for the prevention of endothelial cells and SAT inflammation.

Key structural and functional differences between early and advanced glycation products.

Most of the studies on advanced glycation end products (AGE) have been carried out with uncharacterized mixtures of AGE, so the observed effects cannot be linked to defined structures. Therefore, we analysed the structural differences between glycated human serum albumin (gHSA), a low glycated protein, and AGE-human serum albumin (AGE-HSA), a high glycated protein, and we compared their effects on endothelial functionality. Specifically, we characterized glycation and composition on both early and advanced stage glycation products of gHSA and AGE-HSA by using the MALDI-TOF-mass spectrometry assay. Furthermore, we studied the effects of both types of glycation products on reactive oxygen species (ROS) production and in the expression of vascular and intercellular cell adhesion molecules (VCAM-1 and ICAM-1) on human umbilical endothelial cells (HUVEC). We also measured the adhesion of peripheral blood mononuclear cells (PBMC) to HUVEC. Low concentrations of gHSA enhanced long-lasting ROS production in HUVEC, whereas lower concentrations of AGE-HSA caused the anticipation of the induced extracellular ROS production. Both gHSA and AGE-HSA up-regulated the expression of VCAM-1 and ICAM-1 at mRNA levels. Nevertheless, only AGE-HSA increased protein levels and enhanced the adhesion of PBMC to HUVEC monolayers. Functional differences were observed between gHSA and AGE-HSA, causing the latter an anticipation of the pro-oxidant effects in comparison to gHSA. Moreover, although both molecules induced genetic up-regulation of adhesion molecules in HUVEC, only the high glycated protein functionally increased mononuclear cell adhesion to endothelial monolayers. These observations could have important clinical consequences in the development of diabetic vascular complications.

The downregulation of ΔNp63 in p53-deficient mouse epidermal tumors favors metastatic behavior.

The TP63 gene codes for two major isoform types, TAp63 and ΔNp63, with probable opposite roles in tumorigenesis. The ΔNp63α protein is frequently amplified and overexpressed in different epithelial tumors. Accordingly, it has been considered a potential oncogene. Nonetheless, a possible metastatic suppressor activity has also been suggested based on the experimental observation that its expression is reduced or even absent in advanced invasive tumors. Such metastatic suppressor activities are often related to tumors bearing point mutated TP53 gene. However, its potential roles in TP53-deficient tumors are poorly characterized. Here we show that in spontaneous tumors, induced by the epidermal-specific Trp53 ablation, the reduction of ΔNp63 expression is an early event, whereas it is re-expressed in the lung metastatic lesions. Using knock down and ectopic expression approaches, we show that ΔNp63 expression opposes the epithelial-mesenchymal transition and reduces the metastatic potential of the cells. This process occurs through the modulation of ΔNp63-dependent downstream targets (including transcription factors and microRNAs) likely to play metastatic roles. Further, ΔNp63 also favors the expression of factors involved in iPS reprogramming, thus suggesting that it can also modulate specific stem cell traits in mouse epidermal tumor cells. Overall, our data assign antimetastatic roles to ΔNp63 in the context of p53 deficiency and epidermis.

Glycated human serum albumin induces NF-κB activation and endothelial nitric oxide synthase uncoupling in human umbilical vein endothelial cells.

AIMS: Non-enzymatic glycated proteins could mediate diabetes vascular complications, but the molecular mechanisms are unknown. Our objective was to find new targets involved in the glycated human serum albumin (gHSA)-enhanced extracellular reactive oxygen species (ROS) production in human endothelial cells. METHODS & RESULTS: Some nuclear factors and phosphorylation cascades were analysed. gHSA activated nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), which up-regulated NOX4 and P22PHOX and enhanced ROS production. Pharmacological inhibition of NF-κB reversed gHSA-enhanced NOX4 expression and decreased gHSA-induced ROS production in extra- and intracellular spaces. The inhibition of activator protein-1 (AP-1) induced a rise in NOX4 and P22PHOX subunit expression and a down-regulation of endothelial nitric oxide synthase (eNOS). AP-1 inhibition also enhanced extracellular ROS production in the presence of serum albumin, but not with gHSA. These results were explained by the eNOS uncoupling induced by gHSA, also demonstrated in this study. Phosphatidylinositol 3-kinase or mitogen-activated protein kinase kinase 1/2 did not show to be involved in gHSA-induced ROS production. CONCLUSIONS: All together, the results suggested that gHSA-enhanced ROS production in endothelium is mediated by: 1) NF-κB activation and subsequence up-regulation of NADPH oxidase, 2) eNOS uncoupling. AP-1, although is not directly affected by gHSA, is another target for regulating NADPH oxidase and eNOS expression in endothelial cells.

Advanced glycation end-products as long-term predictors of death and reinfarction after an acute coronary syndrome.

AIM: We evaluated the prognostic value of plasmatic fluorescent advanced glycation end-products (AGE) to predict long-term death and reinfarction in patients with acute coronary syndrome (ACS). MATERIALS & METHODS: A unicenter registry comprising a prospective cohort of 210 ACS patients (47.4% ST-segment elevation myocardial infarction) followed up during 3.1 ± 0.9 years were carried out. RESULTS: Cardiovascular death ratio was 5.7% and 23 patients suffered reinfarction (11.0%). The hazard ratio of the multivariate analysis with respect to death and reinfarction for AGE adjusted by GRACE risk score was 1.011 (1.006-1.016), p < 0.001. CONCLUSION: Fluorescent AGE plasma levels were an independent predictor of death and reinfarction in the long-term follow-up of patients with ACS.

Current status of NADPH oxidase research in cardiovascular pharmacology.

The implications of reactive oxygen species in cardiovascular disease have been known for some decades. Rationally, therapeutic antioxidant strategies combating oxidative stress have been developed, but the results of clinical trials have not been as good as expected. Therefore, to move forward in the design of new therapeutic strategies for cardiovascular disease based on prevention of production of reactive oxygen species, steps must be taken on two fronts, ie, comprehension of reduction-oxidation signaling pathways and the pathophysiologic roles of reactive oxygen species, and development of new, less toxic, and more selective nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitors, to clarify both the role of each NADPH oxidase isoform and their utility in clinical practice. In this review, we analyze the value of NADPH oxidase as a therapeutic target for cardiovascular disease and the old and new pharmacologic agents or strategies to prevent NADPH oxidase activity. Some inhibitors and different direct or indirect approaches are available. Regarding direct NADPH oxidase inhibition, the specificity of NADPH oxidase is the focus of current investigations, whereas the chemical structure-activity relationship studies of known inhibitors have provided pharmacophore models with which to search for new molecules. From a general point of view, small-molecule inhibitors are preferred because of their hydrosolubility and oral bioavailability. However, other possibilities are not closed, with peptide inhibitors or monoclonal antibodies against NADPH oxidase isoforms continuing to be under investigation as well as the ongoing search for naturally occurring compounds. Likewise, some different approaches include inhibition of assembly of the NADPH oxidase complex, subcellular translocation, post-transductional modifications, calcium entry/release, electron transfer, and genetic expression. High-throughput screens for any of these activities could provide new inhibitors. All this knowledge and the research presently underway will likely result in development of new drugs for inhibition of NADPH oxidase and application of therapeutic approaches based on their action, for the treatment of cardiovascular disease in the next few years.

Fluorescent advanced glycation end products and their soluble receptor: the birth of new plasmatic biomarkers for risk stratification of acute coronary syndrome.

OBJECTIVE: Advanced glycation end products (AGEs) have pathophysiological implications in cardiovascular diseases. The aim of our study was to evaluate the prognostic value of fluorescent AGEs and its soluble receptor (sRAGE) in the context of acute coronary syndrome (ACS), both in-hospital phase and follow-up period. METHODS: A prospective clinical study was performed in patients with debut's ACS. The endpoints were the development of cardiac events (cardiac deaths, re-infarction and new-onset heart failure) during in-hospital phase and follow-up period (366 days, inter-quartile range: 273-519 days). 215 consecutive ACS patients admitted to the coronary care unit (62.7±13.0 years, 24.2% female) were included. 47.4% had a diagnosis of ST segment elevation myocardial infarction. AGEs and sRAGE were analysed by fluorescence spectroscopy and competitive ELISA, respectively. Risk scores (GRACE, TIMI, PURSUIT) were calculated retrospectively using prospective data. The complexity of coronary artery disease was evaluated by SYNTAX score. RESULTS: The mean fluorescent AGEs and sRAGE levels were 57.7±45.1 AU and 1045.4±850.0 pg/mL, respectively. 19 patients presented cardiac events during in-hospital phase and 29 during the follow-up. In-hospital cardiac events were significantly associated with higher sRAGE levels (p = 0.001), but not long-term cardiac events (p = 0.365). Regarding fluorescent AGE the opposite happened. After multivariate analysis correcting by gender, left ventricular ejection fraction, glucose levels, haemoglobin, GRACE and SYNTAX scores, sRAGE was significantly associated with in-hospital prognosis, whereas fluorescent AGEs was significantly associated with long-term prognosis. CONCLUSIONS: We conclude that elevated values of sRAGE are associated with worse in-hospital prognosis, whereas high fluorescent AGE levels are associated with more follow-up events.